Sanjiv Sam Gambhir, MD, PhD

All Publications

• Tumor treating fields (TTFields) impairs aberrant glycolysis in glioblastoma as evaluated by [18F]DASA-23, a non-invasive probe of pyruvate kinase M2 (PKM2) expression Neoplasia Patel, C. B., Beinat, C., Xie, Y., Chang, E., Gambhir, S. S. 2021; 23 (1): 58-67

  View details for DOI 10.1016/j.neo.2020.11.003

• Molecular Imaging of Chimeric Antigen Receptor T Cells by ICOS-ImmunoPET. Clinical cancer research : an official journal of the American Association for Cancer Research Simonetta, F., Alam, I. S., Lohmeyer, J. K., Sahaf, B., Good, Z., Chen, W., Xiao, Z., Hirai, T., Scheller, L., Engels, P., Vermesh, O., Robinson, E., Haywood, T., Sathirachinda, A., Baker, J., Malipatlolla, M. B., Schultz, L. M., Spiegel, J. Y., Lee, J. T., Miklos, D. B., Mackall, C. L., Gambhir, S. S., Negrin, R. 2020

  Abstract

  PURPOSE: Immunomonitoring of chimeric antigen receptor (CAR) T cells relies primarily on their quantification in the peripheral blood, which inadequately quantifies their biodistribution and activation status in the tissues. Non-invasive molecular imaging of CAR T cells by positron emission tomography (PET) is a promising approach with the ability to provide spatial, temporal and functional information. Reported strategies rely on the incorporation of reporter transgenes or ex vivo biolabeling, significantly limiting the application of CAR T cell molecular imaging. In the present study, we assessed the ability of antibody-based PET (immunoPET) to non-invasively visualize CAR T cells.EXPERIMENTAL DESIGN: After analyzing human CAR T cells in vitro and ex vivo from patient samples to identify candidate targets for immunoPET, we employed a syngeneic, orthotopic murine tumor model of lymphoma to assess the feasibility of in vivo tracking of CAR T cells by immunoPET using the 89Zr-DFO-anti-ICOS tracer we previously reported.RESULTS: Analysis of human CD19-CAR T cells during activation identified the Inducible T-cell COStimulator (ICOS) as a potential target for immunoPET. In a preclinical tumor model, 89Zr-DFO-ICOS mAb PET-CT imaging detected significantly higher signal in specific bone marrow-containing skeletal sites of CAR T cell treated mice compared with controls. Importantly, administration of ICOS-targeting antibodies at tracer doses did not interfere with CAR T cell persistence and function.CONCLUSIONS: This study highlights the potential of ICOS-immunoPET imaging for monitoring of CAR T cell therapy, a strategy readily applicable to both commercially available and investigational CAR T cells.

  View details for DOI 10.1158/1078-0432.CCR-20-2770

  View details for PubMedID 33087332

• Mitochondrial copper depletion suppresses triple-negative breast cancer in mice. Nature biotechnology Cui, L., Gouw, A. M., LaGory, E. L., Guo, S., Attarwala, N., Tang, Y., Qi, J., Chen, Y., Gao, Z., Casey, K. M., Bazhin, A. A., Chen, M., Hu, L., Xie, J., Fang, M., Zhang, C., Zhu, Q., Wang, Z., Giaccia, A. J., Gambhir, S. S., Zhu, W., Felsher, D. W., Pegram, M. D., Goun, E. A., Le, A., Rao, J. 2020

  Abstract

  Depletion of mitochondrial copper, which shifts metabolism from respiration to glycolysis and reduces energy production, is known to be effective against cancer types that depend on oxidative phosphorylation. However, existing copper chelators are too toxic or ineffective for cancer treatment. Here we develop a safe, mitochondria-targeted, copper-depleting nanoparticle (CDN) and test it against triple-negative breast cancer (TNBC). We show that CDNs decrease oxygen consumption and oxidative phosphorylation, cause a metabolic switch to glycolysis and reduce ATP production in TNBC cells. This energy deficiency, together with compromised mitochondrial membrane potential and elevated oxidative stress, results in apoptosis. CDNs should be less toxic than existing copper chelators because they favorably deprive copper in the mitochondria in cancer cells instead of systemic depletion. Indeed, we demonstrate low toxicity of CDNs in healthy mice. In three mouse models of TNBC, CDN administration inhibits tumor growth and substantially improves survival. The efficacy and safety of CDNs suggest the potential clinical relevance of this approach.

  View details for DOI 10.1038/s41587-020-0707-9

  View details for PubMedID 33077961

• PET reporter gene imaging and ganciclovir-mediated ablation of chimeric antigen receptor T-cells in solid tumors. Cancer research Murty, S., Labanieh, L., Murty, T., Gowrishankar, G., Haywood, T., Alam, I. S., Beinat, C., Robinson, E., Aalipour, A., Klysz, D. D., Cochran, J. R., Majzner, R. G., Mackall, C. L., Gambhir, S. S. 2020

  Abstract

  Imaging strategies to monitor chimeric antigen receptor (CAR) T-cell biodistribution and proliferation harbor the potential to facilitate clinical translation for the treatment of both liquid and solid tumors. Additionally, the potential adverse effects of CAR T-cells highlight the need for mechanisms to modulate CAR T-cell activity. The herpes simplex virus type 1 thymidine kinase (HSV1-tk) gene has previously been translated as a positron emission tomography (PET) reporter gene for imaging of T-cell trafficking in brain tumor patients. The HSV1-TK enzyme can act as a suicide gene of transduced cells through treatment with the prodrug ganciclovir (GCV). Here we report the molecular engineering, imaging, and GCV-mediated destruction of B7H3 CAR T-cells incorporating a mutated version of the HSV1-tk gene (sr39tk) with improved enzymatic activity for GCV. The sr39tk gene did not affect B7H3 CAR T-cell functionality and in vitro and in vivo studies in osteosarcoma models showed no significant effect on B7H3 CAR T-cell antitumor activity. PET/CT imaging with 9-(4-[18F]-fluoro-3-[hydroxymethyl]butyl)guanine [18F]FHBG of B7H3-sr39tk CAR T-cells in an orthotopic model of osteosarcoma revealed tumor homing and systemic immune expansion. Bioluminescence and PET imaging of B7H3-sr39tk CAR T-cells confirmed complete tumor ablation with intraperitoneal GCV administration. This imaging and suicide ablation system can provide insight into CAR T-cell migration and proliferation during clinical trials while serving as a suicide switch to limit potential toxicities.

  View details for DOI 10.1158/0008-5472.CAN-19-3579

  View details for PubMedID 32958548

• Visualization of activated T cells by OX40-immunoPET as a strategy for diagnosis of acute Graft-versus-Host-Disease. Cancer research Alam, I. S., Simonetta, F., Scheller, L., Mayer, A. T., Murty, S., Vermesh, O., Nobashi, T. W., Lohmeyer, J. K., Hirai, T., Baker, J., Lau, K. H., Negrin, R., Gambhir, S. S. 2020

  Abstract

  Graft versus host disease (GvHD) is a major complication of allogeneic hematopoietic cell transplantation (HCT), mediated primarily by donor T cells that become activated and attack host tissues. Non-invasive strategies detecting T cell activation would allow for early diagnosis and possibly more effective management of HCT recipients. Positron emission tomography (PET) imaging is a sensitive and clinically relevant modality ideal for GvHD diagnosis and there is a strong rationale for the use of PET tracers that can monitor T cell activation and expansion with high specificity. The tumor necrosis factor (TNF) receptor superfamily member OX40 (CD134) is a cell surface marker that is highly specific for activated T cells, is upregulated during GvHD, and mediates disease pathogenesis. We recently reported the development of an antibody-based activated T cell imaging agent targeting OX40. In the present study, we visualize the dynamics of OX40 expression in a major histocompatibility complex (MHC)-mismatch mouse model of acute GvHD using OX40-immunoPET. This approach enabled visualization of T cell activation at early stages of disease, prior to overt clinical symptoms with high sensitivity and specificity. This study highlights the potential utility of the OX40 PET imaging as a new strategy for GvHD diagnosis and therapy monitoring.

  View details for DOI 10.1158/0008-5472.CAN-20-1149

  View details for PubMedID 32900772

• Clinical evaluation of (4S)-4-(3-[18F]Fluoropropyl)-L-glutamate (18F-FSPG) for PET/CT imaging in patients with newly diagnosed and recurrent prostate cancer. Clinical cancer research : an official journal of the American Association for Cancer Research Park, S. Y., Na, S. J., Kumar, M., Mosci, C., Wardak, M., Koglin, N., Bullich, S., Mueller, A., Berndt, M., Stephens, A. W., Cho, Y. M., Ahn, H., Chae, S. Y., Kim, H. O., Moon, D. H., Gambhir, S. S., Mittra, E. S. 2020

  Abstract

  PURPOSE: (4S)-4-(3-[18F]Fluoropropyl)-L-glutamic acid (18F-FSPG) is a radiopharmaceutical for PET imaging of system xC - activity, which can be upregulated in prostate cancer. We present data on the first evaluation of newly diagnosed or recurrent prostate cancer patients with this radiopharmaceutical.EXPERIMENTAL DESIGN: Ten primary and ten recurrent prostate cancer patients were enrolled in this prospective multicenter study. After injection of 300 MBq of 18F-FSPG, three whole-body PET/CT scans were obtained. Visual analysis was compared with step-section histopathology when available as well as other imaging studies and clinical outcomes. Metabolic parameters were measured semi-quantitatively. Expression levels of xCT and CD44 were evaluated by immunohistochemistry for patients with available tissue samples.RESULTS: 18F-FSPG PET showed high tumor-to-background ratios with a relatively high tumor detection rate on a per-patient (89%) and per-lobe (87%) basis. The sensitivity was slightly higher with imaging at 105 minutes in comparison to 60 minutes. The SUVmax for cancer was significantly higher than both normal (p < 0.005) and benign pathology (p=0.011), while there was no significant difference between normal and benign pathology (p=0.120). In the setting of recurrence, agreement with standard imaging was demonstrated in 7 of 9 patients (78%) and 13 of 18 lesions (72%), and revealed true local recurrence in a discordant case. 18F-FSPG accumulation showed moderate correlation with CD44 expression.CONCLUSIONS: 18F-FSPG is a promising tumor imaging agent for PET that seems to have favorable biodistribution and high cancer detection rate in patients with prostate cancer. Further studies are warranted to determine the diagnostic value for both initial staging and recurrence, and how it compares to other investigational radiotracers and conventional imaging modalities.

  View details for DOI 10.1158/1078-0432.CCR-20-0644

  View details for PubMedID 32694158

• Synthesis and Characterization of 9-(4-[18F]Fluoro-3-(hydroxymethyl)butyl)-2-(phenylthio)-6-oxopurine as a Novel PET Agent for Mutant Herpes Simplex Virus Type 1 Thymidine Kinase Reporter Gene Imaging. Molecular imaging and biology Fuchigami, T., Haywood, T., Gowrishankar, G., Anders, D., Namavari, M., Wardak, M., Gambhir, S. S. 2020

  Abstract

  PURPOSE: [18F]FHBG has been used as a positron emission tomography (PET) imaging tracer for the monitoring of herpes simplex virus type 1 thymidine kinase (HSV1-tk), a reporter gene for cell and gene therapy in humans. However, this tracer shows inadequate blood-brain barrier (BBB) penetration and, therefore, would be limited for accurate quantification of reporter gene expression in the brain. Here, we report the synthesis and evaluation of 9-(4-[18F]fluoro-3-(hydroxymethyl)butyl)-2(phenylthio)-6-oxopurine ([18F]FHBT) as a new PET tracer for imaging reporter gene expression of HSV1-tk and its mutant HSV1-sr39tk, with the aim of improved BBB penetration.PROCEDURES: [18F]FHBT was prepared by using a tosylate precursor and [18F]KF. The cellular uptake of [18F]FHBT was performed in HSV1-sr39tk-positive (+) or HSV1-sr39tk-negative (-) MDA-MB-231 breast cancer cells. The specificity of [18F]FHBT to assess HSV1-sr39tk expression was evaluated by in vitro blocking studies using 1mM of ganciclovir (GCV). Penetration of [18F]FHBT and [18F]FHBG across the BBB was assessed by dynamic PET imaging studies in normal mice.RESULTS: The tosylate precursor reacted with [18F]KF using Kryptofix2.2.2 followed by deprotection to give [18F]FHBT in 10% radiochemical yield (decay-corrected). The uptake of [18F]FHBT in HSV1-sr39tk (+) cells was significantly higher than that of HSV1-sr39tk (-) cells. In the presence of GCV (1mM), the uptake of [18F]FHBT was significantly decreased, indicating that [18F]FHBT serves as a selective substrate of HSV1-sr39TK. PET images and time-activity curves of [18F]FHBT in the brain regions showed similar initial brain uptakes (~12.75min) as [18F]FHBG (P>0.855). Slower washout of [18F]FHBT was observed at the later time points (17.75 - 57.75 min, P >0.207).CONCLUSIONS: Although [18F]FHBT showed no statistically significant improvement of BBB permeability compared with [18F]FHBG, we have demonstrated that the 2-(phenylthio)-6-oxopurine backbone can serve as a novel scaffold for developing HSV1-tk/HSV1-sr39tk reporter gene imaging agents for additional research in the future.

  View details for DOI 10.1007/s11307-020-01517-5

  View details for PubMedID 32691392

• Isotopically Encoded Nanotags for Multiplexed Ion Beam Imaging. Advanced materials technologies Harmsen, S., Coskun, A. F., Ganesh, S., Nolan, G. P., Gambhir, S. S. 2020; 5 (7)

  Abstract

  High-dimensional profiling of markers and analytes using approaches, such as barcoded fluorescent imaging with repeated labeling and mass cytometry has allowed visualization of biological processes at the single-cell level. To address limitations of sensitivity and mass-channel capacity, a nanobarcoding platform is developed for multiplexed ion beam imaging (MIBI) using secondary ion beam spectrometry that utilizes fabricated isotopically encoded nanotags. Use of combinatorial isotope distributions in 100 nm sized nanotags expands the labeling palette to overcome the spectral bounds of mass channels. As a proof-of-principle, a four-digit (i.e., 0001-1111) barcoding scheme is demonstrated to detect 16 variants of 2H, 19F, 79/81Br, and 127I elemental barcode sets that are encoded in silica nanoparticle matrices. A computational debarcoding method and an automated machine learning analysis approach are developed to extract barcodes for accurate quantification of spatial nanotag distributions in large ion beam imaging areas up to 0.6 mm2. Isotopically encoded nanotags should boost the performance of mass imaging platforms, such as MIBI and other elemental-based bioimaging approaches.

  View details for DOI 10.1002/admt.202000098

  View details for PubMedID 32661501

  View details for PubMedCentralID PMC7357881

• Viral Delivery of CAR Targets to Solid Tumors Enables Effective Cell Therapy. Molecular therapy oncolytics Aalipour, A., Le Boeuf, F., Tang, M., Murty, S., Simonetta, F., Lozano, A. X., Shaffer, T. M., Bell, J. C., Gambhir, S. S. 2020; 17: 232–40

  Abstract

  Chimeric antigen receptor (CAR) Tcell therapy has had limited efficacy for solid tumors, largely due to a lack of selectively and highly expressed surface antigens. To avoid reliance on a tumor's endogenous antigens, here we describe a method of tumor-selective delivery of surface antigens using an oncolytic virus to enable a generalizable CAR Tcell therapy. Using CD19 as our proof of concept, we engineered a thymidine kinase-disrupted vaccinia virus to selectively deliver CD19 to malignant cells, and thus demonstrated potentiation of CD19 CAR Tcell activity against two tumor types invitro. In an immunocompetent model of B16 melanoma, this combination markedly delayed tumor growth and improved median survival compared with antigen-mismatched combinations. We also found that CD19 delivery could improve CAR Tcell activity against tumor cells that express low levels of cognate antigen, suggesting a potential application in counteracting antigen-low escape. This approach highlights the potential of engineering tumors for effective adoptive cell therapy.

  View details for DOI 10.1016/j.omto.2020.03.018

  View details for PubMedID 32346612

• PET imaging of the natural killer cell activation receptor NKp30. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Shaffer, T., Gambhir, S. S., Aalipour, A., Schurch, C. 2020

  Abstract

  Redirecting the immune system in cancer treatment has led to remarkable responses in a subset of patients. Natural killer (NK) cells are innate lymphoid cells being explored as they engage tumor cells in different mechanisms compared to T cells, which could be exploited for treatment of nonresponders to current immunotherapies. NK cell therapies are monitored through measuring peripheral NK cell concentrations or changes in tumor volume over time. The former does not detect NK cells at the tumor site(s), and the latter is inaccurate for immunotherapies because of pseudoprogression. Therefore, new imaging methods are required as companion diagnostics for optimizing immunotherapies. Methods: Here we develop and complete pre-clinical in vivo validation of two antibody-based PET probes specific for NKp30, an activation natural cytotoxicity receptor expressed by human NK cells. Quantitative, multicolor flow cytometry during a variety of NK cell activation conditions was completed on primary human NK cells and the NK92MI cell line. Human renal cell carcinoma (RCC) tumors were stained for the NK cell receptors CD56, NKp30, and NKp46 to determine expression on tumor-infiltrating NK cells. An NKp30 antibody was radiolabeled with 64Cu or 89Zr and evaluated in subcutaneous xenografts and adoptive cell transfer mouse models. Results: Quantitative flow cytometry showed consistent expression of the NKp30 receptor during different activation conditions. NKp30 and NKp46 costained in RCC samples, demonstrating the expression of these receptors on tumor-infiltrating NK cells in human tumors, while tumor cells in one RCC sample expressed the peripheral NK marker CD56. Both PET tracers showed high stability and specificity in vitro and in vivo. Notably, 89Zr- NKp30Ab had higher on-target contrast compared to 64Cu-NKpAb at their respective terminal time points. 64Cu-NKp30Ab delineated NK cell trafficking to the liver and spleen in an adoptive cell transfer model. Conclusion: The consistent expression of NKp30 on NK cells makes it an attractive target for quantitative imaging. Immunofluorescence staining on human RCC samples demonstrated the advantages of NKp30 targeting versus the traditional CD56 for detection of tumor infiltrating NK cells. This work advances PET imaging of NK cells and supports the translation of imaging agents for immunotherapy monitoring.

  View details for DOI 10.2967/jnumed.119.233163

  View details for PubMedID 32532927

• The Project Baseline Health Study: a step towards a broader mission to map human health NPJ DIGITAL MEDICINE Arges, K., Assimes, T., Bajaj, V., Balu, S., Bashir, M. R., Beskow, L., Blanco, R., Califf, R., Campbell, P., Carin, L., Christian, V., Cousins, S., Das, M., Dockery, M., Douglas, P. S., Dunham, A., Eckstrand, J., Fleischmann, D., Ford, E., Fraulo, E., French, J., Gambhir, S. S., Ginsburg, G. S., Green, R. C., Haddad, F., Hernandez, A., Hernandez, J., Huang, E. S., Jaffe, G., King, D., Koweek, L. H., Langlotz, C., Liao, Y. J., Mahaffey, K. W., Marcom, K., Marks, W. J., Maron, D., McCabe, R., McCall, S., McCue, R., Mega, J., Miller, D., Muhlbaier, L. H., Munshi, R., Newby, L., Pak-Harvey, E., Patrick-Lake, B., Pencina, M., Peterson, E. D., Rodriguez, F., Shore, S., Shah, S., Shipes, S., Sledge, G., Spielman, S., Spitler, R., Schaack, T., Swamy, G., Willemink, M. J., Wong, C. A. 2020; 3 (1): 84

  Abstract

  The Project Baseline Health Study (PBHS) was launched to map human health through a comprehensive understanding of both the health of an individual and how it relates to the broader population. The study will contribute to the creation of a biomedical information system that accounts for the highly complex interplay of biological, behavioral, environmental, and social systems. The PBHS is a prospective, multicenter, longitudinal cohort study that aims to enroll thousands of participants with diverse backgrounds who are representative of the entire health spectrum. Enrolled participants will be evaluated serially using clinical, molecular, imaging, sensor, self-reported, behavioral, psychological, environmental, and other health-related measurements. An initial deeply phenotyped cohort will inform the development of a large, expanded virtual cohort. The PBHS will contribute to precision health and medicine by integrating state of the art testing, longitudinal monitoring and participant engagement, and by contributing to the development of an improved platform for data sharing and analysis.

  View details for DOI 10.1038/s41746-020-0290-y

  View details for Web of Science ID 000538242900001

  View details for PubMedID 32550652

  View details for PubMedCentralID PMC7275087

• Molecular Imaging of Infective Endocarditis With 6''-[18F]Fluoromaltotriose Positron Emission Tomography-Computed Tomography. Circulation Wardak, M., Gowrishankar, G., Zhao, X., Liu, Y., Chang, E., Namavari, M., Haywood, T., Gabr, M. T., Neofytou, E., Chour, T., Qin, X., Vilches-Moure, J. G., Hardy, J., Contag, C. H., McConnell, M. V., Wu, J. C., Gambhir, S. S. 2020; 141 (21): 1729–31

  View details for DOI 10.1161/CIRCULATIONAHA.119.043924

  View details for PubMedID 32453662

• Isotopically Encoded Nanotags for Multiplexed Ion Beam Imaging ADVANCED MATERIALS TECHNOLOGIES Harmsen, S., Coskun, A. F., Ganesh, S., Nolan, G. P., Gambhir, S. S. 2020

  View details for DOI 10.1002/admt.202000098

  View details for Web of Science ID 000530513000001

• A pilot study of F-18-FSPG SiPM-based PET/CT in patients referred for exclusion of active cardiac sarcoidosis and negative or non-diagnostic F-18-FDG PET/CT Duan, H., Hatami, N., Baratto, L., Davidzon, G., Aparici, C., Gambhir, S., Koglin, N., Witteles, R., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2020

  View details for Web of Science ID 000568290501539

• Imaging activated immune response following therapeutic vaccination in an orthotopic glioma model with Zr-89-DFO-OX40 mAb PET Nobashi, T., Mayer, A., Xiao, Z., Chan, C., Chaney, A., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2020

  View details for Web of Science ID 000568290500003

• Non-Invasive Photoacoustic Imaging of In Vivo Mice with Erythrocyte Derived Optical Nanoparticles to Detect CAD/MI. Scientific reports Liu, Y., Hanley, T., Chen, H., Long, S. R., Gambhir, S. S., Cheng, Z., Wu, J. C., Fakhri, G. E., Anvari, B., Zaman, R. T. 2020; 10 (1): 5983

  Abstract

  Coronary artery disease (CAD) causes mortality and morbidity worldwide. We used near-infrared erythrocyte-derived transducers (NETs), a contrast agent, in combination with a photoacoustic imaging system to identify the locations of atherosclerotic lesions and occlusion due to myocardial-infarction (MI). NETs (90nm diameter) were fabricated from hemoglobin-depleted mice erythrocyte-ghosts and doped with Indocyanine Green (ICG). Ten weeks old male C57BL/6 mice (n=9) underwent left anterior descending (LAD) coronary artery ligation to mimic vulnerable atherosclerotic plaques and their rupture leading to MI. 150L of NETs (20M ICG,) was IV injected via tail vein 1-hour prior to photoacoustic (PA) and fluorescence in vivo imaging by exciting NETs at 800nm and 650nm, respectively. These results were verified with histochemical analysis. We observed 256-fold higher PA signal from the accumulated NETs in the coronary artery above the ligation. Fluorescence signals were detected in LAD coronary, thymus, and liver. Similar signals were observed when the chest was cut open. Atherosclerotic lesions exhibited inflammatory cells. Liver demonstrated normal portal tract, with no parenchymal necrosis, inflammation, fibrosis, or other pathologic changes, suggesting biocompatibility of NETs. Non-invasively detecting atherosclerotic plaques and stenosis using NETs may lay a groundwork for future clinical detection and improving CAD risk assessment.

  View details for DOI 10.1038/s41598-020-62868-1

  View details for PubMedID 32249814

• Positron emission tomography (PET) imaging of the natural killer (NK) cell activation receptor NKp30. Shaffer, T. M., Aalipour, A., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2020: 102–3

  View details for Web of Science ID 000522837200165

• Integrating genomic features for non-invasive early lung cancer detection. Nature Chabon, J. J., Hamilton, E. G., Kurtz, D. M., Esfahani, M. S., Moding, E. J., Stehr, H., Schroers-Martin, J., Nabet, B. Y., Chen, B., Chaudhuri, A. A., Liu, C. L., Hui, A. B., Jin, M. C., Azad, T. D., Almanza, D., Jeon, Y. J., Nesselbush, M. C., Co Ting Keh, L., Bonilla, R. F., Yoo, C. H., Ko, R. B., Chen, E. L., Merriott, D. J., Massion, P. P., Mansfield, A. S., Jen, J., Ren, H. Z., Lin, S. H., Costantino, C. L., Burr, R., Tibshirani, R., Gambhir, S. S., Berry, G. J., Jensen, K. C., West, R. B., Neal, J. W., Wakelee, H. A., Loo, B. W., Kunder, C. A., Leung, A. N., Lui, N. S., Berry, M. F., Shrager, J. B., Nair, V. S., Haber, D. A., Sequist, L. V., Alizadeh, A. A., Diehn, M. 2020; 580 (7802): 245-251

  Abstract

  Radiologic screening of high-risk adults reduces lung-cancer-related mortality1,2; however, a small minority of eligible individuals undergo such screening in the United States3,4. The availability of blood-based tests could increase screening uptake. Here we introduce improvements to cancer personalized profiling by deep sequencing (CAPP-Seq)5, a method for the analysis of circulating tumour DNA (ctDNA), to better facilitate screening applications. We show that, although levels are very low in early-stage lung cancers, ctDNA is present prior to treatment in most patients and its presence is strongly prognostic. We also find that the majority of somatic mutations in the cell-free DNA (cfDNA) of patients with lung cancer and of risk-matched controls reflect clonal haematopoiesis and are non-recurrent. Compared with tumour-derived mutations, clonal haematopoiesis mutations occur on longer cfDNA fragments and lack mutational signatures that are associated with tobacco smoking. Integrating these findings with other molecular features, we develop and prospectively validate a machine-learning method termed 'lung cancer likelihood in plasma' (Lung-CLiP), which can robustly discriminate early-stage lung cancer patients from risk-matched controls. This approach achieves performance similar to that of tumour-informed ctDNA detection and enables tuning of assay specificity in order to facilitate distinct clinical applications. Our findings establish the potential of cfDNA for lung cancer screening and highlight the importance of risk-matching cases and controls in cfDNA-based screening studies.

  View details for DOI 10.1038/s41586-020-2140-0

  View details for PubMedID 32269342

• ICOS is an indicator of T cell-mediated response to cancer immunotherapy. Cancer research Xiao, Z., Mayer, A. T., Nobashi, T. W., Gambhir, S. S. 2020

  Abstract

  Immunotherapy is innovating clinical cancer management. Nevertheless, only a small fraction of patients benefit from current immunotherapies. To improve clinical management of cancer immunotherapy, it is critical to develop strategies for response monitoring and prediction. In this study, we describe Inducible T cell Costimulator (ICOS) as a conserved mediator of immune response across multiple therapy strategies. ICOS expression was evaluated by flow cytometry, 89Zr-DFO-ICOS mAb PET/CT imaging was performed on Lewis lung cancer models treated with different immunotherapy strategies, and the change in tumor volume was used as a read-out for therapeutic response. ImmunoPET imaging of ICOS enabled sensitive and specific detection of activated T cells and early benchmarking of immune response. A STING agonist was identified as a promising therapeutic approach in this manner. The STING agonist generated significantly stronger immune responses as measured by ICOS ImmunoPET and delayed tumor growth compared to PD-1 checkpoint blockade. More importantly, ICOS ImmunoPET enabled early and robust prediction of therapeutic response across multiple treatment regimens. This data shows that ICOS is an indicator of T cell-mediated immune response and suggests ICOS ImmunoPET as a promising strategy for monitoring, comparing, and predicting immunotherapy success in cancer.

  View details for DOI 10.1158/0008-5472.CAN-19-3265

  View details for PubMedID 32156777

• SP94-Targeted Triblock Copolymer Nanoparticle Delivers Thymidine Kinase-p53-Nitroreductase Triple Therapeutic Gene and Restores Anticancer Function against Hepatocellular Carcinoma in Vivo. ACS applied materials & interfaces Sukumar, U. K., Rajendran, J. C., Gambhir, S. S., Massoud, T. F., Paulmurugan, R. 2020

  Abstract

  Gene-directed enzyme-prodrug therapy (GDEPT) is a promising approach for cancer therapy, but it suffers from poor targeted delivery in vivo. Polyethylenimine (PEI) is a cationic polymer efficient in delivering negatively charged nucleic acids across cell membranes; however, it is highly toxic in vivo. Hence, we efficiently reduced PEI toxicity without compromising its transfection efficiency by conjugating it with poly(d,l-lactic-co-glycolic acid) (PLGA) and poly(ethylene glycol) (PEG) as triblock copolymers through a multistep synthetic process. The synthesized nanoparticles showed efficient delivery of loaded nucleic acids to tumor cells in vitro and in vivo in mice. We used this nanoparticle to deliver a rationally engineered thymidine kinase (TK)-p53-nitroreductase (NTR) triple therapeutic gene against hepatocellular carcinoma (HCC), where p53 tumor suppressor gene is mutated in more than 85% of cancers. TK-p53-NTR triple gene therapy restores p53 function and potentiates cancer cell response to delivered prodrugs (ganciclovir (GCV) and CB1954). We used SP94 peptide-functionalized PLGA-PEG-PEI nanoparticles for the optimal delivery of TK-p53-NTR therapeutic gene in vivo. The nanoparticles prepared from the conjugated polymer showed high loading efficiency for the DNA and markedly enhanced TK-NTR-mediated gene therapy upon the simultaneous coexpression of p53 by the concurrent rescue of the endogenous apoptotic pathway in HCC cells of both p53-mutant and wild-type phenotypes in vitro. In vivo delivery of TK-p53-NTR genes by SP94-targeted PLGA-PEG-PEI NP in mice resulted in a strong expression of suicide genes selectively in tumors, and subsequent administration of GCV and CB1954 led to a decline in tumor growth, and established a superior therapeutic outcome against HCC. We demonstrate a highly efficient approach that exogenously supplements p53 to enable synergy with the outcome of TK-NTR suicide gene therapy against HCC.

  View details for DOI 10.1021/acsami.9b20071

  View details for PubMedID 32048820

• Radiotheranostics: a roadmap for future development LANCET ONCOLOGY Herrmann, K., Schwaiger, M., Lewis, J. S., Solomon, S. B., McNeil, B. J., Baumann, M., Gambhir, S. S., Hricak, H., Weissleder, R. 2020; 21 (3): E146–E156

  View details for Web of Science ID 000518470100019

• Evaluation of Glycolytic Response to Multiple Classes of Anti-glioblastoma Drugs by Noninvasive Measurement of Pyruvate Kinase M2 Using [F-18]DASA-23 MOLECULAR IMAGING AND BIOLOGY Beinat, C., Patel, C. B., Xie, Y., Gambhir, S. S. 2020; 22 (1): 124–33

  View details for DOI 10.1007/s11307-019-01353-2

  View details for Web of Science ID 000513265700018

• Carbon-coated FeCo nanoparticles as sensitive magnetic-particle-imaging tracers with photothermal and magnetothermal properties. Nature biomedical engineering Song, G. n., Kenney, M. n., Chen, Y. S., Zheng, X. n., Deng, Y. n., Chen, Z. n., Wang, S. X., Gambhir, S. S., Dai, H. n., Rao, J. n. 2020

  Abstract

  The low magnetic saturation of iron oxide nanoparticles, which are developed primarily as contrast agents for magnetic resonance imaging, limits the sensitivity of their detection using magnetic particle imaging (MPI). Here, we show that FeCo nanoparticles that have a core diameter of 10 nm and bear a graphitic carbon shell decorated with poly(ethylene glycol) provide an MPI signal intensity that is sixfold and fifteenfold higher than the signals from the superparamagnetic iron oxide tracers VivoTrax and Feraheme, respectively, at the same molar concentration of iron. We also show that the nanoparticles have photothermal and magnetothermal properties and can therefore be used for tumour ablation in mice, and that they have high optical absorbance in a broad near-infrared region spectral range (wavelength, 700-1,200 nm), making them suitable as tracers for photoacoustic imaging. As sensitive multifunctional and multimodal imaging tracers, carbon-coated FeCo nanoparticles may confer advantages in cancer imaging and hyperthermia therapy.

  View details for DOI 10.1038/s41551-019-0506-0

  View details for PubMedID 32015409

• Whole-body tracking of single cells via positron emission tomography. Nature biomedical engineering Jung, K. O., Kim, T. J., Yu, J. H., Rhee, S. n., Zhao, W. n., Ha, B. n., Red-Horse, K. n., Gambhir, S. S., Pratx, G. n. 2020

  Abstract

  In vivo molecular imaging can measure the average kinetics and movement routes of injected cells through the body. However, owing to non-specific accumulation of the contrast agent and its efflux from the cells, most of these imaging methods inaccurately estimate the distribution of the cells. Here, we show that single human breast cancer cells loaded with mesoporous silica nanoparticles concentrating the 68Ga radioisotope and injected into immunodeficient mice can be tracked in real time from the pattern of annihilation photons detected using positron emission tomography, with respect to anatomical landmarks derived from X-ray computed tomography. The cells travelled at an average velocity of 50 mm s-1 and arrested in the lungs 2-3 s after tail-vein injection into the mice, which is consistent with the blood-flow rate. Single-cell tracking could be used to determine the kinetics of cell trafficking and arrest during the earliest phase of the metastatic cascade, the trafficking of immune cells during cancer immunotherapy and the distribution of cells after transplantation.

  View details for DOI 10.1038/s41551-020-0570-5

  View details for PubMedID 32541917

• Initial evaluation of (4S)-4-(3-[18F]fluoropropyl)-L-glutamate (FSPG) PET/CT imaging in patients with head and neck cancer, colorectal cancer, or non-Hodgkin lymphoma. EJNMMI research Park, S. Y., Mosci, C. n., Kumar, M. n., Wardak, M. n., Koglin, N. n., Bullich, S. n., Mueller, A. n., Berndt, M. n., Stephens, A. W., Chin, F. T., Gambhir, S. S., Mittra, E. S. 2020; 10 (1): 100

  Abstract

  (4S)-4-(3-[18F]Fluoropropyl)-L-glutamic acid ([18F]FSPG) measures system xC- transporter activity and shows promise for oncologic imaging. We present data on tumor uptake of this radiopharmaceutical in human subjects with head and neck cancer (HNC), colorectal cancer (CRC), and non-Hodgkin lymphoma (NHL).A total of 15 subjects with HNC (n = 5), CRC (n = 5), or NHL (n = 5) were recruited (mean age 66.2 years, range 44-87 years). 301.4 ± 28.1 MBq (8.1 ± 0.8 mCi) of [18F]FSPG was given intravenously to each subject, and 3 PET/CT scans were obtained 0-2 h post-injection. All subjects also had a positive [18F]FDG PET/CT scan within 1 month prior to the [18F]FSPG PET scan. Semi-quantitative and visual comparisons of the [18F]FSPG and [18F]FDG scans were performed.[18F]FSPG showed strong uptake in all but one HNC subject. The lack of surrounding brain uptake facilitated tumor delineation in the HNC patients. [18F]FSPG also showed tumor uptake in all CRC subjects, but variable uptake in the NHL subjects. While the absolute [18F]FDG SUV values were comparable or higher than [18F]FSPG, the tumor-to-background SUV ratios were greater with [18F]FSPG than [18F]FDG.[18F]FSPG PET/CT showed promising results across 15 subjects with 3 different cancer types. Concordant visualization was mostly observed between [18F]FSPG and [18F]FDG PET/CT images, with some inter- and intra-individual uptake variability potentially reflecting differences in tumor biology. The tumor-to-background ratios were greater with [18F]FSPG than [18F]FDG in the cancer types evaluated. Future studies based on larger numbers of subjects and those with a wider array of primary and recurrent or metastatic tumors are planned to further evaluate the utility of this novel tracer.

  View details for DOI 10.1186/s13550-020-00678-2

  View details for PubMedID 32857284

• Multispectral Photoacoustic Assessment of Thyroid Cancer Nodules In Vivo Kim, J., Park, B., Ha, J., Steinberg, I., Park, E., Choi, W., Hooper, S., Gambhir, S., Lim, D., Kim, C., Oraevsky, A. A., Wang, L. V. SPIE-INT SOC OPTICAL ENGINEERING. 2020

  View details for DOI 10.1117/12.2546616

  View details for Web of Science ID 000558347500001

• Molecular Imaging of Chimeric Antigen Receptor T Cells by ICOS-ImmunoPET Clinical cancer research: an official journal of the American Association for Cancer Research Alam*, I. S., Simonetta*, F. 2020: 1058–68

  Abstract

  Immunomonitoring of chimeric antigen receptor (CAR) T cells relies primarily on their quantification in the peripheral blood, which inadequately quantifies their biodistribution and activation status in the tissues. Noninvasive molecular imaging of CAR T cells by PET is a promising approach with the ability to provide spatial, temporal, and functional information. Reported strategies rely on the incorporation of reporter transgenes or ex vivo biolabeling, significantly limiting the application of CAR T-cell molecular imaging. In this study, we assessed the ability of antibody-based PET (immunoPET) to noninvasively visualize CAR T cells.After analyzing human CAR T cells in vitro and ex vivo from patient samples to identify candidate targets for immunoPET, we employed a syngeneic, orthotopic murine tumor model of lymphoma to assess the feasibility of in vivo tracking of CAR T cells by immunoPET using the 89Zr-DFO-anti-ICOS tracer, which we have previously reported.Analysis of human CD19-CAR T cells during activation identified the Inducible T-cell COStimulator (ICOS) as a potential target for immunoPET. In a preclinical tumor model, 89Zr-DFO-ICOS mAb PET-CT imaging detected significantly higher signal in specific bone marrow-containing skeletal sites of CAR T-cell-treated mice compared with controls. Importantly, administration of ICOS-targeting antibodies at tracer doses did not interfere with CAR T-cell persistence and function.This study highlights the potential of ICOS-immunoPET imaging for monitoring of CAR T-cell therapy, a strategy readily applicable to both commercially available and investigational CAR T cells.See related commentary by Volpe et al., p. 911.

  View details for DOI 10.1158/1078-0432.CCR-20-2770

  View details for PubMedCentralID PMC7887027

• A mountable toilet system for personalized health monitoring via the analysis of excreta. Nature biomedical engineering Park, S. M., Won, D. D., Lee, B. J., Escobedo, D. n., Esteva, A. n., Aalipour, A. n., Ge, T. J., Kim, J. H., Suh, S. n., Choi, E. H., Lozano, A. X., Yao, C. n., Bodapati, S. n., Achterberg, F. B., Kim, J. n., Park, H. n., Choi, Y. n., Kim, W. J., Yu, J. H., Bhatt, A. M., Lee, J. K., Spitler, R. n., Wang, S. X., Gambhir, S. S. 2020

  Abstract

  Technologies for the longitudinal monitoring of a person's health are poorly integrated with clinical workflows, and have rarely produced actionable biometric data for healthcare providers. Here, we describe easily deployable hardware and software for the long-term analysis of a user's excreta through data collection and models of human health. The 'smart' toilet, which is self-contained and operates autonomously by leveraging pressure and motion sensors, analyses the user's urine using a standard-of-care colorimetric assay that traces red-green-blue values from images of urinalysis strips, calculates the flow rate and volume of urine using computer vision as a uroflowmeter, and classifies stool according to the Bristol stool form scale using deep learning, with performance that is comparable to the performance of trained medical personnel. Each user of the toilet is identified through their fingerprint and the distinctive features of their anoderm, and the data are securely stored and analysed in an encrypted cloud server. The toilet may find uses in the screening, diagnosis and longitudinal monitoring of specific patient populations.

  View details for DOI 10.1038/s41551-020-0534-9

  View details for PubMedID 32251391

• Radiotheranostics: a roadmap for future development. The Lancet. Oncology Herrmann, K. n., Schwaiger, M. n., Lewis, J. S., Solomon, S. B., McNeil, B. J., Baumann, M. n., Gambhir, S. S., Hricak, H. n., Weissleder, R. n. 2020; 21 (3): e146–e156

  Abstract

  Radiotheranostics, injectable radiopharmaceuticals with antitumour effects, have seen rapid development over the past decade. Although some formulations are already approved for human use, more radiopharmaceuticals will enter clinical practice in the next 5 years, potentially introducing new therapeutic choices for patients. Despite these advances, several challenges remain, including logistics, supply chain, regulatory issues, and education and training. By highlighting active developments in the field, this Review aims to alert practitioners to the value of radiotheranostics and to outline a roadmap for future development. Multidisciplinary approaches in clinical trial design and therapeutic administration will become essential to the continued progress of this evolving therapeutic approach.

  View details for DOI 10.1016/S1470-2045(19)30821-6

  View details for PubMedID 32135118

• New synthesis of 6''-[18 F]fluoromaltotriose for positron emission tomography (PET) imaging of bacterial infection. Journal of labelled compounds & radiopharmaceuticals Gabr, M. T., Haywood, T. n., Gowrishankar, G. n., Srinivasan, A. n., Gambhir, S. S. 2020

  Abstract

  6''-[18 F]fluoromaltotriose is a positron emission tomography (PET) tracer that can differentiate between bacterial infection and inflammation in vivo. Bacteria-specific uptake of 6''-[18 F]fluoromaltotriose is attributed to the targeting of maltodextrin transporter in bacteria that is absent in mammalian cells. Herein, we report a new synthesis of 6''-[18 F]fluoromaltotriose as a key step for its clinical translation. In comparison to the previously reported synthesis, the new synthesis features unambiguous assignment of the fluorine-18 position on the maltotriose unit. The new method utilizes direct fluorination of 2'',3'',4''-tri-O-acetyl-6''-O-trifyl-α-D-glucopyranosyl-(1-4)-O-2',3',6'-tri-O-acetyl-α-D-glucopyranosyl-(1-4)-1,2,3,6-tetra-O-acetyl-D-glucopyranose followed by basic hydrolysis. Radiolabeling of the new maltotriose triflate precursor proceeds using a single HPLC purification step, which results in shorter reaction time in comparison to the previously reported synthesis. Successful synthesis of 6''-[18 F]fluoromaltotriose has been achieved in 3.5 ± 0.3% radiochemical yield (decay corrected, n=7) and radiochemical purity above 95%. The efficient radiosynthesis of 6''-[18 F]fluoromaltotriose would be critical in advancing this PET tracer into clinical trials for imaging bacterial infections.

  View details for DOI 10.1002/jlcr.3868

  View details for PubMedID 32602175

• Intravital imaging reveals synergistic effect of CAR T-cells and radiation therapy in a preclinical immunocompetent glioblastoma model Oncoimmunology Murty, S., Haile, S. T., Beinat, C., Aalipour, A., Alam, I. S., Murty, T., Shaffer, T. M., Patel, C. B., Graves, E. E., Mackall, C. L., Gambhir, S. S. 2020; 9 (1)

  View details for DOI 10.1080/2162402X.2020.1757360

• Reconstructed Apoptotic Bodies as Targeted "Nano Decoys" to Treat Intracellular Bacterial Infections within Macrophages and Cancer Cells. ACS nano Bose, R. J., Tharmalingam, N. n., Garcia Marques, F. J., Sukumar, U. K., Natarajan, A. n., Zeng, Y. n., Robinson, E. n., Bermudez, A. n., Chang, E. n., Habte, F. n., Pitteri, S. J., McCarthy, J. R., Gambhir, S. S., Massoud, T. F., Mylonakis, E. n., Paulmurugan, R. n. 2020

  Abstract

  Staphylococcus aureus (S. aureus) is a highly pathogenic facultative anaerobe that in some instances resides as an intracellular bacterium within macrophages and cancer cells. This pathogen can establish secondary infection foci, resulting in recurrent systemic infections that are difficult to treat using systemic antibiotics. Here, we use reconstructed apoptotic bodies (ReApoBds) derived from cancer cells as "nano decoys" to deliver vancomycin intracellularly to kill S. aureus by targeting inherent "eat me" signaling of ApoBds. We prepared ReApoBds from different cancer cells (SKBR3, MDA-MB-231, HepG2, U87-MG, and LN229) and used them for vancomycin delivery. Physicochemical characterization showed ReApoBds size ranges from 80 to 150 nm and vancomycin encapsulation efficiency of 60 ± 2.56%. We demonstrate that the loaded vancomycin was able to kill intracellular S. aureus efficiently in an in vitro model of S. aureus infected RAW-264.7 macrophage cells, and U87-MG (p53-wt) and LN229 (p53-mt) cancer cells, compared to free-vancomycin treatment (P < 0.001). The vancomycin loaded ReApoBds treatment in S. aureus infected macrophages showed a two-log-order higher CFU reduction than the free-vancomycin treatment group. In vivo studies revealed that ReApoBds can specifically target macrophages and cancer cells. Vancomycin loaded ReApoBds have the potential to kill intracellular S. aureus infection in vivo in macrophages and cancer cells.

  View details for DOI 10.1021/acsnano.0c00921

  View details for PubMedID 32347709

• Low-frequency ultrasound-mediated cytokine transfection enhances T cell recruitment at local and distant tumor sites. Proceedings of the National Academy of Sciences of the United States of America Ilovitsh, T. n., Feng, Y. n., Foiret, J. n., Kheirolomoom, A. n., Zhang, H. n., Ingham, E. S., Ilovitsh, A. n., Tumbale, S. K., Fite, B. Z., Wu, B. n., Raie, M. N., Zhang, N. n., Kare, A. J., Chavez, M. n., Qi, L. S., Pelled, G. n., Gazit, D. n., Vermesh, O. n., Steinberg, I. n., Gambhir, S. S., Ferrara, K. W. 2020

  Abstract

  Robust cytotoxic T cell infiltration has proven to be difficult to achieve in solid tumors. We set out to develop a flexible protocol to efficiently transfect tumor and stromal cells to produce immune-activating cytokines, and thus enhance T cell infiltration while debulking tumor mass. By combining ultrasound with tumor-targeted microbubbles, membrane pores are created and facilitate a controllable and local transfection. Here, we applied a substantially lower transmission frequency (250 kHz) than applied previously. The resulting microbubble oscillation was significantly enhanced, reaching an effective expansion ratio of 35 for a peak negative pressure of 500 kPa in vitro. Combining low-frequency ultrasound with tumor-targeted microbubbles and a DNA plasmid construct, 20% of tumor cells remained viable, and ∼20% of these remaining cells were transfected with a reporter gene both in vitro and in vivo. The majority of cells transfected in vivo were mucin 1+/CD45- tumor cells. Tumor and stromal cells were then transfected with plasmid DNA encoding IFN-β, producing 150 pg/106 cells in vitro, a 150-fold increase compared to no-ultrasound or no-plasmid controls and a 50-fold increase compared to treatment with targeted microbubbles and ultrasound (without IFN-β). This enhancement in secretion exceeds previously reported fourfold to fivefold increases with other in vitro treatments. Combined with intraperitoneal administration of checkpoint inhibition, a single application of IFN-β plasmid transfection reduced tumor growth in vivo and recruited efficacious immune cells at both the local and distant tumor sites.

  View details for DOI 10.1073/pnas.1914906117

  View details for PubMedID 32430322

• Discovery and Optimization of Small-Molecule Ligands for V-Domain Ig Suppressor of T-Cell Activation (VISTA). Journal of the American Chemical Society Gabr, M. T., Gambhir, S. S. 2020

  Abstract

  V-domain Ig suppressor of T-cell activation (VISTA) is an immune checkpoint that affects the ability of T-cells to attack tumors. A FRET-based high throughput screening identified NSC622608 as the first small-molecule ligand for VISTA. Investigation of the interaction of NSC622608 with VISTA using STD NMR and molecular modeling enabled the identification of a potential binding site in VISTA for NSC622608. Screening NSC622608 against a library of single-point VISTA mutants revealed the key residues in VISTA interacting with NSC622608. Further structural optimization resulted in a lead with submicromolar VISTA binding affinity. The lead compound blocked VISTA signaling in vitro, enhanced T-cell proliferation, and restored T-cell activation in the presence of VISTA-expressing cancer cell lines. This work would enable future development of small molecules targeting VISTA as immunomodulators and imaging probes.

  View details for DOI 10.1021/jacs.0c07276

  View details for PubMedID 32894020

• Author Correction: Non-Invasive Photoacoustic Imaging of In Vivo Mice with Erythrocyte Derived Optical Nanoparticles to Detect CAD/MI. Scientific reports Liu, Y. n., Hanley, T. n., Chen, H. n., Long, S. R., Gambhir, S. S., Cheng, Z. n., Wu, J. C., Fakhri, G. E., Anvari, B. n., Zaman, R. T. 2020; 10 (1): 19102

  Abstract

  An amendment to this paper has been published and can be accessed via a link at the top of the paper.

  View details for DOI 10.1038/s41598-020-75966-x

  View details for PubMedID 33127974

• Plasmonic and Electrostatic Interactions Enable Uniformly Enhanced Liquid Bacterial Surface-Enhanced Raman Scattering (SERS). Nano letters Tadesse, L. F., Ho, C. S., Chen, D. H., Arami, H. n., Banaei, N. n., Gambhir, S. S., Jeffrey, S. S., Saleh, A. A., Dionne, J. n. 2020

  Abstract

  Surface-enhanced Raman spectroscopy (SERS) is a promising cellular identification and drug susceptibility testing platform, provided it can be performed in a controlled liquid environment that maintains cell viability. We investigate bacterial liquid-SERS, studying plasmonic and electrostatic interactions between gold nanorods and bacteria that enable uniformly enhanced SERS. We synthesize five nanorod sizes with longitudinal plasmon resonances ranging from 670 to 860 nm and characterize SERS signatures of Gram-negative Escherichia coli and Serratia marcescens and Gram-positive Staphylococcus aureus and Staphylococcus epidermidis bacteria in water. Varying the concentration of bacteria and nanorods, we achieve large-area SERS enhancement that is independent of nanorod resonance and bacteria type; however, bacteria with higher surface charge density exhibit significantly higher SERS signal. Using cryo-electron microscopy and zeta potential measurements, we show that the higher signal results from attraction between positively charged nanorods and negatively charged bacteria. Our robust liquid-SERS measurements provide a foundation for bacterial identification and drug testing in biological fluids.

  View details for DOI 10.1021/acs.nanolett.0c03189

  View details for PubMedID 32914987

• Toward the Clinical Development and Validation of a Thy1-Targeted Ultrasound Contrast Agent for the Early Detection of Pancreatic Ductal Adenocarcinoma. Investigative radiology Bam, R. n., Daryaei, I. n., Abou-Elkacem, L. n., Vilches-Moure, J. G., Meuillet, E. J., Lutz, A. n., Marinelli, E. R., Unger, E. C., Gambhir, S. S., Paulmurugan, R. n. 2020

  Abstract

  Early detection of pancreatic ductal adenocarcinoma (PDAC) represents the most significant step toward the treatment of this aggressive lethal disease. Previously, we engineered a preclinical Thy1-targeted microbubble (MBThy1) contrast agent that specifically recognizes Thy1 antigen overexpressed in the vasculature of murine PDAC tissues by ultrasound (US) imaging. In this study, we adopted a single-chain variable fragment (scFv) site-specific bioconjugation approach to construct clinically translatable MBThy1-scFv and test for its efficacy in vivo in murine PDAC imaging, and functionally evaluated the binding specificity of scFv ligand to human Thy1 in patient PDAC tissues ex vivo.We recombinantly expressed the Thy1-scFv with a carboxy-terminus cysteine residue to facilitate its thioether conjugation to the PEGylated MBs presenting with maleimide functional groups. After the scFv-MB conjugations, we tested binding activity of the MBThy1-scFv to MS1 cells overexpressing human Thy1 (MS1Thy1) under liquid shear stress conditions in vitro using a flow chamber setup at 0.6 mL/min flow rate, corresponding to a wall shear stress rate of 100 seconds, similar to that in tumor capillaries. For in vivo Thy1 US molecular imaging, MBThy1-scFv was tested in the transgenic mouse model (C57BL/6J - Pdx1-Cre; KRas; Ink4a/Arf) of PDAC and in control mice (C57BL/6J) with L-arginine-induced pancreatitis or normal pancreas. To facilitate its clinical feasibility, we further produced Thy1-scFv without the bacterial fusion tags and confirmed its recognition of human Thy1 in cell lines by flow cytometry and in patient PDAC frozen tissue sections of different clinical grades by immunofluorescence staining.Under shear stress flow conditions in vitro, MBThy1-scFv bound to MS1Thy1 cells at significantly higher numbers (3.0 ± 0.8 MB/cell; P < 0.01) compared with MBNontargeted (0.5 ± 0.5 MB/cell). In vivo, MBThy1-scFv (5.3 ± 1.9 arbitrary units [a.u.]) but not the MBNontargeted (1.2 ± 1.0 a.u.) produced high US molecular imaging signal (4.4-fold vs MBNontargeted; n = 8; P < 0.01) in the transgenic mice with spontaneous PDAC tumors (2-6 mm). Imaging signal from mice with L-arginine-induced pancreatitis (n = 8) or normal pancreas (n = 3) were not significantly different between the two MB constructs and were significantly lower than PDAC Thy1 molecular signal. Clinical-grade scFv conjugated to Alexa Fluor 647 dye recognized MS1Thy1 cells but not the parental wild-type cells as evaluated by flow cytometry. More importantly, scFv showed highly specific binding to VEGFR2-positive vasculature and fibroblast-like stromal components surrounding the ducts of human PDAC tissues as evaluated by confocal microscopy.Our findings summarize the development and validation of a clinically relevant Thy1-targeted US contrast agent for the early detection of human PDAC by US molecular imaging.

  View details for DOI 10.1097/RLI.0000000000000697

  View details for PubMedID 32569010

• Reduction Triggered In Situ Polymerization in Living Mice. Journal of the American Chemical Society Cui, L. n., Vivona, S. n., Smith, B. R., Kothapalli, S. R., Liu, J. n., Ma, X. n., Chen, Z. n., Taylor, M. n., Kierstead, P. H., Fréchet, J. M., Gambhir, S. S., Rao, J. n. 2020

  Abstract

  "Smart" biomaterials that are responsive to physiological or biochemical stimuli have found many biomedical applications for tissue engineering, therapeutics, and molecular imaging. In this work, we describe in situ polymerization of activatable biorthogonal small molecules in response to a reducing environment change in vivo. We designed a carbohydrate linker- and cyanobenzothiazole-cysteine condensation reaction-based small molecule scaffold that can undergo rapid condensation reaction upon physiochemical changes (such as a reducing environment) to form polymers (pseudopolysaccharide). The fluorescent and photoacoustic properties of a fluorophore-tagged condensation scaffold before and after the transformation have been examined with a dual-modality optical imaging method. These results confirmed the in situ polymerization of this probe after both local and systemic administration in living mice.

  View details for DOI 10.1021/jacs.0c07594

  View details for PubMedID 32804495

• Human biodistribution and radiation dosimetry of [18F]DASA-23, a PET probe targeting pyruvate kinase M2. European journal of nuclear medicine and molecular imaging Beinat, C. n., Patel, C. B., Haywood, T. n., Shen, B. n., Naya, L. n., Gandhi, H. n., Holley, D. n., Khalighi, M. n., Iagaru, A. n., Davidzon, G. n., Gambhir, S. S. 2020

  Abstract

  To assess the safety, biodistribution, and radiation dosimetry of the novel positron emission tomography (PET) radiopharmaceutical 1-((2-fluoro-6-[[18F]]fluorophenyl)sulfonyl)-4-((4-methoxyphenyl)sulfonyl)piperazine ([18F]DASA-23) in healthy volunteers.We recruited 5 healthy volunteers who provided a written informed consent. Volunteers were injected with 295.0 ± 8.2 MBq of [18F]DASA-23 intravenously. Immediately following injection, a dynamic scan of the brain was acquired for 15 min. This was followed by serial whole-body PET/MRI scans acquired up to 3 h post-injection. Blood samples were collected at regular intervals, and vital signs monitored pre- and post-radiotracer administration. Regions of interest were drawn around multiple organs, time-activity curves were calculated, and organ uptake and dosimetry were estimated with OLINDA/EXM (version 1.1) software.All subjects tolerated the PET/MRI examination, without adverse reactions to [18F]DASA-23. [18F]DASA-23 passively crossed the blood-brain barrier, followed by rapid clearance from the brain. High accumulation of [18F]DASA-23 was noted in organs such as the gallbladder, liver, small intestine, and urinary bladder, suggesting hepatobiliary and urinary clearance. The effective dose of [18F]DASA-23 was 23.5 ± 5.8 μSv/MBq.We successfully completed a pilot first-in-human study of [18F]DASA-23. Our results indicate that [18F]DASA-23 can be used safely in humans to evaluate pyruvate kinase M2 levels. Ongoing studies are evaluating the ability of [18F]DASA-23 to visualize intracranial malignancies, NCT03539731.ClinicalTrials.gov, NCT03539731 (registered 28 May 2018).

  View details for DOI 10.1007/s00259-020-04687-0

  View details for PubMedID 31938892

• Trop2 is a driver of metastatic prostate cancer with neuroendocrine phenotype via PARP1. Proceedings of the National Academy of Sciences of the United States of America Hsu, E. C., Rice, M. A., Bermudez, A. n., Marques, F. J., Aslan, M. n., Liu, S. n., Ghoochani, A. n., Zhang, C. A., Chen, Y. S., Zlitni, A. n., Kumar, S. n., Nolley, R. n., Habte, F. n., Shen, M. n., Koul, K. n., Peehl, D. M., Zoubeidi, A. n., Gambhir, S. S., Kunder, C. A., Pitteri, S. J., Brooks, J. D., Stoyanova, T. n. 2020

  Abstract

  Resistance to androgen deprivation therapy, or castration-resistant prostate cancer (CRPC), is often accompanied by metastasis and is currently the ultimate cause of prostate cancer-associated deaths in men. Recently, secondary hormonal therapies have led to an increase of neuroendocrine prostate cancer (NEPC), a highly aggressive variant of CRPC. Here, we identify that high levels of cell surface receptor Trop2 are predictive of recurrence of localized prostate cancer. Moreover, Trop2 is significantly elevated in CRPC and NEPC, drives prostate cancer growth, and induces neuroendocrine phenotype. Overexpression of Trop2 induces tumor growth and metastasis while loss of Trop2 suppresses these abilities in vivo. Trop2-driven NEPC displays a significant up-regulation of PARP1, and PARP inhibitors significantly delay tumor growth and metastatic colonization and reverse neuroendocrine features in Trop2-driven NEPC. Our findings establish Trop2 as a driver and therapeutic target for metastatic prostate cancer with neuroendocrine phenotype and suggest that high Trop2 levels could identify cancers that are sensitive to Trop2-targeting therapies and PARP1 inhibition.

  View details for DOI 10.1073/pnas.1905384117

  View details for PubMedID 31932422

• Two Patient Studies of a Companion Diagnostic Immuno-Positron Emission Tomography (PET) Tracer for Measuring Human CA6 Expression in Cancer for Antibody Drug Conjugate (ADC) Therapy. Molecular imaging Natarajan, A., Srinivas, S. M., Azevedo, C., Greene, L., Bauchet, A., Jouannot, E., Lacoste-Bourgeacq, A., Guizon, I., Cohen, P., Naneix, A., Ilovich, O., Cisneros, J., Rupanarayan, K., Chin, F. T., Iagaru, A., Dirbas, F. M., Karam, A., Gambhir, S. S. 2020; 19: 1536012120939398

  Abstract

  An antigen binding fragment (BFab) derived from a tumor-associated mucin 1-sialoglycotope antigen (CA6) targeting antibody (huDS6) was engineered. We synthesized a companion diagnostic positron emission tomography (PET) tracer by radiolabeling BFab with [64Cu] to measure CA6 expression on cancer tissues prior to anti-human CA6 (huDS6-DM4 antibody-drug conjugate) therapy for ovarian and breast cancer patients. After chemotherapy, the ovarian patient received PET scan with 18F-2-fluoro-2-deoxyglucose ([18F]FDG: 10 mCi), followed by [64Cu]-DOTA-BFab ([64Cu]BFab; 5.5 mCi) 1 week later for PET scanning of CA6 expression and subsequent surgery. The breast cancer patient was treated with chemotherapy before primary tumor resection and subsequent [18F]FDG-PET scan. 4 weeks later the patient received of [64Cu]BFab (11.7 mCi) for CA6 PET scan. Whole body [18F]FDG-PET of the breast cancer patient indicated FDG-avid tumor metastases to the liver, bilateral hila and thoracic spine, but no uptake was observed for the ovarian patient. Each patient was also imaged by PET/CT with [64Cu]BFab at 1 and 24 hours after tracer administration. The [64Cu]BFab tracer was well tolerated by both patients without adverse effects, and no significant tracer uptake was observed in both patients. Immunohistochemistry (IHC) data indicated CA6 expressions were weak to intermediate and matched with the [64Cu]BFab-PET signals.

  View details for DOI 10.1177/1536012120939398

  View details for PubMedID 33104454

• Publisher Correction: A mountable toilet system for personalized health monitoring via the analysis of excreta. Nature biomedical engineering Park, S. M., Won, D. D., Lee, B. J., Escobedo, D. n., Esteva, A. n., Aalipour, A. n., Ge, T. J., Kim, J. H., Suh, S. n., Choi, E. H., Lozano, A. X., Yao, C. n., Bodapati, S. n., Achterberg, F. B., Kim, J. n., Park, H. n., Choi, Y. n., Kim, W. J., Yu, J. H., Bhatt, A. M., Lee, J. K., Spitler, R. n., Wang, S. X., Gambhir, S. S. 2020

  Abstract

  An amendment to this paper has been published and can be accessed via a link at the top of the paper.

  View details for DOI 10.1038/s41551-020-0562-5

  View details for PubMedID 32382068

• Maltotriose-based probes for fluorescence and photoacoustic imaging of bacterial infections. Nature communications Zlitni, A. n., Gowrishankar, G. n., Steinberg, I. n., Haywood, T. n., Sam Gambhir, S. n. 2020; 11 (1): 1250

  Abstract

  Currently, there are no non-invasive tools to accurately diagnose wound and surgical site infections before they become systemic or cause significant anatomical damage. Fluorescence and photoacoustic imaging are cost-effective imaging modalities that can be used to noninvasively diagnose bacterial infections when paired with a molecularly targeted infection imaging agent. Here, we develop a fluorescent derivative of maltotriose (Cy7-1-maltotriose), which is shown to be taken up in a variety of gram-positive and gram-negative bacterial strains in vitro. In vivo fluorescence and photoacoustic imaging studies highlight the ability of this probe to detect infection, assess infection burden, and visualize the effectiveness of antibiotic treatment in E. coli-induced myositis and a clinically relevant S. aureus wound infection murine model. In addition, we show that maltotriose is an ideal scaffold for infection imaging agents encompassing better pharmacokinetic properties and in vivo stability than other maltodextrins (e.g. maltohexose).

  View details for DOI 10.1038/s41467-020-14985-8

  View details for PubMedID 32144257

• Biodegradable fluorescent nanoparticles for endoscopic detection of colorectal carcinogenesis. Advanced functional materials Rogalla, S., Flisikowski, K., Gorpas, D., Mayer, A. T., Flisikowska, T., Mandella, M. J., Ma, X., Casey, K. M., Felt, S. A., Saur, D., Ntziachristos, V., Schnieke, A., Contag, C. H., Gambhir, S. S., Harmsen, S. 2019; 29 (51)

  Abstract

  Early and comprehensive endoscopic detection of colonic dysplasia - the most clinically significant precursor lesion to colorectal adenocarcinoma - provides an opportunity for timely, minimally-invasive intervention to prevent malignant transformation. Here, the development and evaluation of biodegradable near-infrared fluorescent silica nanoparticles (FSN) is described that have the potential to improve adenoma detection during fluorescence-assisted white-light colonoscopic surveillance in rodent and human-scale models of colorectal carcinogenesis. FSNs are biodegradable (t1/2 of 2.7 weeks), well-tolerated, and enable detection and delineation of adenomas as small as 0.5 mm2 with high tumor-to-background ratios. Furthermore, in the human-scale, APC 1311/+ porcine model, the clinical feasibility and benefit of using FSN-guided detection of colorectal adenomas using video-rate fluorescence-assisted white-light endoscopy is demonstrated. Since nanoparticles of similar size (e.g., 100-150-nm) or composition (i.e., silica, silica/gold hybrid) have already been successfully translated to the clinic, and, clinical fluorescent/white light endoscopy systems are becoming more readily available, there is a viable path towards clinical translation of the proposed strategy for early colorectal cancer detection and prevention in high-risk patients.

  View details for DOI 10.1002/adfm.201904992

  View details for PubMedID 33041743

  View details for PubMedCentralID PMC7546531

• Biodegradable Fluorescent Nanoparticles for Endoscopic Detection of Colorectal Carcinogenesis ADVANCED FUNCTIONAL MATERIALS Rogalla, S., Flisikowski, K., Gorpas, D., Mayer, A. T., Flisikowska, T., Mandella, M. J., Ma, X., Casey, K. M., Felt, S. A., Saur, D., Ntziachristos, V., Schnieke, A., Contag, C. H., Gambhir, S. S., Harmsen, S. 2019; 29 (51)

  View details for DOI 10.1002/adfm.201904992

  View details for Web of Science ID 000516572400007

• TUMOR TREATING FIELDS LEADS TO CHANGES IN MEMBRANE PERMEABILITY AND INCREASED PENETRATION BY ANTI-GLIOMA DRUGS Chang, E., Patel, C., Beinat, C., Young, C., Flores, T., Zeng, Y., Joubert, L., Arami, H., Natarajan, A., Sinclair, R., Gambhir, S. OXFORD UNIV PRESS INC. 2019: 93

  View details for Web of Science ID 000509478702048

• EVALUATION OF [18F]DASA-23 FOR NON-INVASIVE MEASUREMENT OF ABERRANTLY EXPRESSED PYRUVATE KINASE M2 IN GLIOMA: FIRST-IN-HUMAN STUDY Patel, C., Beinat, C., Haywood, T., Murty, S., Xie, Y., Recht, L., Nagpal, S., Thomas, R., Khalighi, M., Gandhi, H., Holley, D., Gambhir, S. OXFORD UNIV PRESS INC. 2019: 169

  View details for Web of Science ID 000509478704010

• Engineering of a novel subnanomolar affinity fibronectin III domain binder targeting human programmed death-ligand 1. Protein engineering, design & selection : PEDS Ramakrishnan, S., Natarajan, A., Chan, C. T., Panesar, P. S., Gambhir, S. S. 2019

  Abstract

  The programmed death-ligand 1 (PD-L1) is a major checkpoint protein that helps cancer cells evade the immune system. A non-invasive imaging agent with rapid clearance rate would be an ideal tool to predict and monitor the efficacy of anti-PD-L1 therapy. The aim of this research was to engineer a subnanomolar, high-affinity fibronectin type 3 domain (FN3)-based small binder targeted against human PD-L1 (hPD-L1) present on tumor cells. A naive yeast G4 library containing the FN3 gene with three binding loop sequences was used to isolate high-affinity binders targeted to purified full-length hPD-L1. The selected binder clones displayed several mutations in the loop regions of the FN3 domain. One unique clone (FN3hPD-L1-01) with a 6x His-tag at the C-terminus had a protein yield of >5mg/L and a protein mass of 12kDa. In vitro binding assays on six different human cancer cell lines (MDA-MB-231, DLD1, U87, 293T, Raji and Jurkat) and murine CT26 colon carcinoma cells stably expressing hPD-L1 showed that CT26/hPD-L1 cells had the highest expression of hPD-L1 in both basal and IFN-gamma-induced states, with a binding affinity of 2.38±0.26nM for FN3hPD-L1-01. The binding ability of FN3hPD-L1-01 was further confirmed by immunofluorescence staining on ex vivo CT26/hPD-L1 tumors sections. The FN3hPD-L1-01 binder represents a novel, small, high-affinity binder for imaging hPD-L1 expression on tumor cells and would aid in earlier imaging of tumors. Future clinical validation studies of the labeled FN3hPD-L1 binder(s) have the potential to monitor immune checkpoint inhibitors therapy and predict responders.

  View details for DOI 10.1093/protein/gzz030

  View details for PubMedID 31612217

• Evaluation of integrin alphavbeta6 cystine knot PET tracers to detect cancer and idiopathic pulmonary fibrosis. Nature communications Kimura, R. H., Wang, L., Shen, B., Huo, L., Tummers, W., Filipp, F. V., Guo, H. H., Haywood, T., Abou-Elkacem, L., Baratto, L., Habte, F., Devulapally, R., Witney, T. H., Cheng, Y., Tikole, S., Chakraborti, S., Nix, J., Bonagura, C. A., Hatami, N., Mooney, J. J., Desai, T., Turner, S., Gaster, R. S., Otte, A., Visser, B. C., Poultsides, G. A., Norton, J., Park, W., Stolowitz, M., Lau, K., Yang, E., Natarajan, A., Ilovich, O., Srinivas, S., Srinivasan, A., Paulmurugan, R., Willmann, J., Chin, F. T., Cheng, Z., Iagaru, A., Li, F., Gambhir, S. S. 2019; 10 (1): 4673

  Abstract

  Advances in precision molecular imaging promise to transform our ability to detect, diagnose and treat disease. Here, we describe the engineering and validation of a new cystine knot peptide (knottin) that selectively recognizes human integrin alphavbeta6 with single-digit nanomolar affinity. We solve its 3D structure by NMR and x-ray crystallography and validate leads with 3 different radiolabels in pre-clinical models of cancer. We evaluate the lead tracer's safety, biodistribution and pharmacokinetics in healthy human volunteers, and show its ability to detect multiple cancers (pancreatic, cervical and lung) in patients at two study locations. Additionally, we demonstrate that the knottin PET tracers can also detect fibrotic lung disease in idiopathic pulmonary fibrosis patients. Our results indicate that these cystine knot PET tracers may have potential utility in multiple disease states that are associated with upregulation of integrin alphavbeta6.

  View details for DOI 10.1038/s41467-019-11863-w

  View details for PubMedID 31611594

• How to Prevent a Leaky Pipeline in Academic Radiology: Insights From a Faculty Survey JOURNAL OF THE AMERICAN COLLEGE OF RADIOLOGY Daldrup-Link, H., Villavasso, K., Zhao, Q., Lu, Y., Ranieri, A., Simard, C., Gambhir, S. 2019; 16 (9): 1220–24

  View details for DOI 10.1016/j.jacr.2019.04.008

  View details for Web of Science ID 000486132000023

• In Vivo Translation of the CIRPI System: Revealing Molecular Pathology of Rabbit Aortic Atherosclerotic Plaques JOURNAL OF NUCLEAR MEDICINE Zaman, R. T., Yousefi, S., Chibana, H., Ikeno, F., Long, S. R., Gambhir, S. S., Chin, F. T., McConnell, M. V., Xing, L., Yeung, A. 2019; 60 (9): 1308–16

  View details for DOI 10.2967/jnumed.118.222471

  View details for Web of Science ID 000484372100026

• Simultaneous transrectal ultrasound and photoacoustic human prostate imaging. Science translational medicine Kothapalli, S., Sonn, G. A., Choe, J. W., Nikoozadeh, A., Bhuyan, A., Park, K. K., Cristman, P., Fan, R., Moini, A., Lee, B. C., Wu, J., Carver, T. E., Trivedi, D., Shiiba, L., Steinberg, I., Huland, D. M., Rasmussen, M. F., Liao, J. C., Brooks, J. D., Khuri-Yakub, P. T., Gambhir, S. S. 2019; 11 (507)

  Abstract

  Imaging technologies that simultaneously provide anatomical, functional, and molecular information are emerging as an attractive choice for disease screening and management. Since the 1980s, transrectal ultrasound (TRUS) has been routinely used to visualize prostatic anatomy and guide needle biopsy, despite limited specificity. Photoacoustic imaging (PAI) provides functional and molecular information at ultrasonic resolution based on optical absorption. Combining the strengths of TRUS and PAI approaches, we report the development and bench-to-bedside translation of an integrated TRUS and photoacoustic (TRUSPA) device. TRUSPA uses a miniaturized capacitive micromachined ultrasonic transducer array for simultaneous imaging of anatomical and molecular optical contrasts [intrinsic: hemoglobin; extrinsic: intravenous indocyanine green (ICG)] of the human prostate. Hemoglobin absorption mapped vascularity of the prostate and surroundings, whereas ICG absorption enhanced the intraprostatic photoacoustic contrast. Future work using the TRUSPA device for biomarker-specific molecular imaging may enable a fundamentally new approach to prostate cancer diagnosis, prognostication, and therapeutic monitoring.

  View details for DOI 10.1126/scitranslmed.aav2169

  View details for PubMedID 31462508

• Microvesicle-mediated delivery of minicircle DNA results in effective gene-directed enzyme prodrug cancer therapy. Molecular cancer therapeutics Kanada, M., Kim, B. D., Hardy, J. W., Bachmann, M. H., Bernard, M. P., Perez, G. I., Zarea, A. A., Ge, T. J., Withrow, A., Ibrahim, S. A., Toomajian, V., Gambhir, S. S., Paulmurugan, R., Contag, C. H., Ronald, J. A. 2019

  Abstract

  An emerging approach for cancer treatment employs the use of extracellular vesicles (EVs), specifically exosomes and microvesicles, as delivery vehicles. We previously demonstrated that microvesicles can functionally deliver plasmid DNA to cells and showed that plasmid size and sequence, in part, determine the delivery efficiency. In this study, delivery vehicles comprised of microvesicles loaded with engineered minicircle (MC) DNA that encodes prodrug converting enzymes were developed as a cancer therapy in mammary carcinoma models. We demonstrated that MCs can be loaded into shed microvesicles with greater efficiency than their parental plasmid counterparts and that microvesicle-mediated MC delivery led to significantly higher and more prolonged transgene expression in recipient cells than microvesicles loaded with the parental plasmid. Microvesicles loaded with MCs encoding a thymidine kinase (TK)/nitroreductase (NTR) fusion protein produced prolonged TK-NTR expression in mammary carcinoma cells. In vivo delivery of TK-NTR and administration of prodrugs led to the effective killing of both targeted cells and surrounding tumor cells via TK-NTR-mediated conversion of co-delivered prodrugs into active cytotoxic agents. In vivo evaluation of the bystander effect in mouse models demonstrated that for effective therapy, at least 1% of tumor cells need to be delivered with TK-NTR-encoding MCs. These results suggest that MC delivery via microvesicles can mediate gene transfer to an extent that enables effective prodrug conversion and tumor cell death such that it comprises a promising approach to cancer therapy.

  View details for DOI 10.1158/1535-7163.MCT-19-0299

  View details for PubMedID 31451563

• Continuous-Wave Coherent Raman Spectroscopy via Plasmonic Enhancement. Scientific reports Monfared, Y. E., Shaffer, T. M., Gambhir, S. S., Hewitt, K. C. 2019; 9 (1): 12092

  Abstract

  In this paper, we report a successful combination of stimulated Raman spectroscopy (SRS) and surface-enhanced Raman scattering (SERS) using cw laser sources and gold/silica nanoparticles with embedded reporter molecules. We describe the preparation method for our gold/silica nanoparticles as well as the effect of probe wavelength, pump and probe power, polarization and sample concentration on the cwSESRS signal. Altogether, a stable ~12 orders of magnitude enhancement in the stimulated Raman signal is achieved because of the amplification of both pump and probe beams, leading to the detection of pico-molar nanoparticle concentrations, comparable to those of SERS. The coherent Raman spectra matches the incoherent conventional Raman spectra of the reporter molecules. Unlike conventional incoherent SERS this approach generates a coherent stimulated signal of microwatt intensities, opening the field to applications requiring a coherent beam, such as Molecular Holography.

  View details for DOI 10.1038/s41598-019-48573-8

  View details for PubMedID 31431666

• Ultrasound/microbubble-mediated targeted delivery of anticancer microRNA-loaded nanoparticles to deep tissues in pigs. Journal of controlled release : official journal of the Controlled Release Society Di Ianni, T., Bose, R. J., Sukumar, U. K., Bachawal, S., Wang, H., Telichko, A., Herickhoff, C., Robinson, E., Baker, S., Vilches-Moure, J. G., Felt, S. A., Gambhir, S. S., Paulmurugan, R., Dahl, J. D. 2019

  Abstract

  In this study, we designed and validated a platform for ultrasound and microbubble-mediated delivery of FDA-approved pegylated poly lactic-co-glycolic acid (PLGA) nanoparticles loaded with anticancer microRNAs (miRNAs) to deep tissues in a pig model. Small RNAs have been shown to reprogram tumor cells and sensitize them to clinically used chemotherapy. To overcome their short intravascular circulation half-life and achieve controlled and sustained release into tumor cells, anticancer miRNAs need to be encapsulated into nanocarriers. Focused ultrasound combined with gas-filled microbubbles provides a noninvasive way to improve the permeability of tumor vasculature and increase the delivery efficiency of drug-loaded particles. A single handheld, curvilinear ultrasound array was used in this study for image-guided therapy with clinical-grade SonoVue contrast agent. First, we validated the platform on phantoms to optimize the microbubble cavitation dose based on acoustic parameters, including peak negative pressure, pulse length, and pulse repetition frequency. We then tested the system in vivo by delivering PLGA nanoparticles co-loaded with antisense-miRNA-21 and antisense-miRNA-10b to pig liver and kidney. Enhanced miRNA delivery was observed (1.9- to 3.7-fold increase) as a result of the ultrasound treatment compared to untreated control regions. Additionally, we used highly fluorescent semiconducting polymer nanoparticles to visually assess nanoparticle extravasation. Fluorescent microscopy suggested the presence of nanoparticles in the extravascular compartment. Hematoxylin and eosin staining of treated tissues did not reveal tissue damage. The results presented in this manuscript suggest that the proposed platform may be used to safely and noninvasively enhance the delivery of miRNA-loaded nanoparticles to target regions in deep organs in large animal models.

  View details for DOI 10.1016/j.jconrel.2019.07.024

  View details for PubMedID 31326463

• Intranasal delivery of targeted polyfunctional gold-iron oxide nanoparticles loaded with therapeutic microRNAs for combined theranostic multimodality imaging and presensitization of glioblastoma to temozolomide. Biomaterials Sukumar, U. K., Bose, R. J., Malhotra, M., Babikir, H. A., Afjei, R., Robinson, E., Zeng, Y., Chang, E., Habte, F., Sinclair, R., Gambhir, S. S., Massoud, T. F., Paulmurugan, R. 2019; 218: 119342

  Abstract

  The prognosis for glioblastoma (GBM) remains depressingly low. The biological barriers of the brain present a major challenge to achieving adequate drug concentrations for GBM therapy. To address this, we explore the potential of the nose-to-brain direct transport pathway to bypass the blood-brain barrier, and to enable targeted delivery of theranostic polyfunctional gold-iron oxide nanoparticles (polyGIONs) surface loaded with therapeutic miRNAs (miR-100 and antimiR-21) to GBMs in mice. These nanoformulations would thus allow presensitization of GBM cells to the systemically delivered chemotherapy drug temozolomide (TMZ), as well as in vivo multimodality molecular and anatomic imaging of nanoparticle delivery, trafficking, and treatment effects. First, we synthesized GIONs coated with beta-cyclodextrin-chitosan (CD-CS) hybrid polymer, and co-loaded with miR-100 and antimiR-21. Then we decorated their surface with PEG-T7 peptide using CD-adamantane host-guest chemistry. The resultant polyGIONs showed efficient miRNA loading with enhanced serum stability. We characterized them for particle size, PDI, polymer functionalization, charge and release using dynamic light scattering analysis, TEM and qRT-PCR. For in vivo intranasal delivery, we used U87-MG GBM cell-derived orthotopic xenograft models in mice. Intranasal delivery resulted in efficient accumulation of Cy5-miRNAs in mice treated with T7-targeted polyGIONs, as demonstrated by in vivo optical fluorescence and MR imaging. We measured the therapeutic response of these FLUC-EGFP labelled U87-MG GBMs using bioluminescence imaging. Overall, there was a significant increase in survival of mice co-treated with T7-polyGIONs loaded with miR-100/antimiR-21 plus systemic TMZ, compared to the untreated control group, or the animals receiving non-targeted polyGIONs-miR-100/antimiR-21, or TMZ alone. Once translated clinically, this novel theranostic nanoformulation and its associated intranasal delivery strategy will have a strong potential to potentiate the effects of TMZ treatment in GBM patients.

  View details for DOI 10.1016/j.biomaterials.2019.119342

  View details for PubMedID 31326657

• Author Correction: Miniature gold nanorods for photoacoustic molecular imaging in the second near-infrared optical window. Nature nanotechnology Chen, Y., Zhao, Y., Yoon, S. J., Gambhir, S. S., Emelianov, S. 2019

  Abstract

  An amendment to this paper has been published and can be accessed via a link at the top of the paper.

  View details for DOI 10.1038/s41565-019-0522-y

  View details for PubMedID 31289408

• Tumor treating fields increases membrane permeability in glioblastoma cells Chang, E., Patel, C., Pohling, C., Young, C., Song, J., Flores, T. A., Zeng, Y., Joubert, L., Arami, H., Natarajan, A., Sinclair, R., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2019

  View details for DOI 10.1158/1538-7445.AM2019-250

  View details for Web of Science ID 000488129901424

• Detection of visually occult metastatic lymph nodes using molecularly targeted fluorescent imaging during surgical resection of pancreatic cancer HPB Tummers, W. S., Miller, S. E., Teraphongphom, N. T., van den Berg, N. S., Hasan, A., Longacre, T. A., Fisher, G. A., Bonsing, B. A., Vahrmeijer, A. L., Gambhir, S. S., Swijnenburg, R., Rosenthal, E. L., Poultsides, G. A. 2019; 21 (7): 883–90

  View details for DOI 10.1016/j.hpb.2018.11.008

  View details for Web of Science ID 000475487400014

• Simultaneous PET/MRI in the Evaluation of Breast and Prostate Cancer Using Combined Na[18F] F and [18F]FDG: a Focus on Skeletal Lesions. Molecular imaging and biology : MIB : the official publication of the Academy of Molecular Imaging Sonni, I., Minamimoto, R., Baratto, L., Gambhir, S. S., Loening, A. M., Vasanawala, S. S., Iagaru, A. 2019

  Abstract

  PURPOSE: The purpose of this study is to prospectively evaluate the performance of sodium 18F]fluoride (Na[18F]F)/2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) simultaneous time-of-flight enabled positron emission tomography (PET)/magnetic resonance imaging (MRI) for the detection of skeletal metastases in selected patients with advanced breast and prostate cancers.PROCEDURE: The institutional review board approved this HIPAA-compliant protocol. Written informed consent was obtained from each patient. A total of 74 patients (23 women and 51 men with breast and prostate cancer, respectively) referred for standard-of-care whole-body bone scintigraphy (WBBS) were enrolled in this prospective study. All patients underwent a [99mTc]methyldiphosphonate ([99mTc]MDP) WBBS followed by Na[18F]F/[18F]FDG PET/MRI. Lesions detected by each imaging modality were tabulated and a lesion-based and patient-based analysis was conducted.RESULTS: On a patient-based analysis, [99mTc]MDP WBBS identified skeletal lesions in 37 patients and PET/MRI in 45 patients. On a lesion-based analysis, WBBS identified a total of 81 skeletal lesions, whereas PET/MRI identified 140 lesions. Additionally, PET/MRI showed extra-skeletal lesions in 19 patients, including lymph nodes (16), prostate (4) lung (3), and liver (2) lesions.CONCLUSIONS: The ability of Na[18F]F/[18F]FDG PET/MRI to identify more skeletal lesions than 99mTc-MDP WBBS and to additionally identify extra-skeletal disease may be beneficial for patient care and represent an alternative to the single modalities performed separately. Na[18F]F/[18F]FDG PET/MRI is a promising approach for evaluation of skeletal and extra-skeletal lesions in a selected population of breast and prostate cancer patients.

  View details for DOI 10.1007/s11307-019-01392-9

  View details for PubMedID 31236756

• Proceedings: Pathways for Successful Translation of New Imaging Agents and Modalities-Phase III Studies JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Shankar, L. K., Rosenthal, E., Warram, J. M., Ghesani, M., Hope, T. A., Jacobs, P. M., Jacobson, G. B., Wilson, T., Siegel, B. A. 2019; 60 (6): 736–44

  View details for DOI 10.2967/jnumed.118.219824

  View details for Web of Science ID 000470084400016

• Photoacoustic clinical imaging. Photoacoustics Steinberg, I., Huland, D. M., Vermesh, O., Frostig, H. E., Tummers, W. S., Gambhir, S. S. 2019; 14: 77–98

  Abstract

  Photoacoustic is an emerging biomedical imaging modality, which allows imaging optical absorbers in the tissue by acoustic detectors (light in - sound out). Such a technique has an immense potential for clinical translation since it allows high resolution, sufficient imaging depth, with diverse endogenous and exogenous contrast, and is free from ionizing radiation. In recent years, tremendous developments in both the instrumentation and imaging agents have been achieved. These opened avenues for clinical imaging of various sites allowed applications such as brain functional imaging, breast cancer screening, diagnosis of psoriasis and skin lesions, biopsy and surgery guidance, the guidance of tumor therapies at the reproductive and urological systems, as well as imaging tumor metastases at the sentinel lymph nodes. Here we survey the various clinical and pre-clinical literature and discuss the potential applications and hurdles that still need to be overcome.

  View details for DOI 10.1016/j.pacs.2019.05.001

  View details for PubMedID 31293884

• Evaluation of [18F]DASA-23 for non-invasive measurement of aberrantly expressed pyruvate kinase M2 in glioblastoma: preclinical and first in human studies Beinat, C., Patel, C., Haywood, T., Murty, S., Alam, I., Xie, Y., Gandhi, H., Holley, D., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2019

  View details for Web of Science ID 000473116800052

• Miniature gold nanorods for photoacoustic molecular imaging in the second near-infrared optical window NATURE NANOTECHNOLOGY Chen, Y., Zhao, Y., Yoon, S., Gambhir, S., Emelianov, S. 2019; 14 (5): 465–72

  View details for DOI 10.1038/s41565-019-0392-3

  View details for Web of Science ID 000467053100023

• [F-18]-SuPAR: A Radiofluorinated Probe for Noninvasive Imaging of DNA Damage-Dependent Poly(ADP-ribose) Polymerase Activity BIOCONJUGATE CHEMISTRY Shuhendler, A. J., Cui, L., Chen, Z., Shen, B., Chen, M., James, M. L., Witney, T. H., Bazalova-Carter, M., Gambhir, S. S., Chin, F. T., Graves, E. E., Rao, J. 2019; 30 (5): 1331–42

  Abstract

  Poly(ADP ribose) polymerase (PARP) enzymes generate poly(ADP ribose) post-translational modifications on target proteins for an array of functions centering on DNA and cell stress. PARP isoforms 1 and 2 are critically charged with the surveillance of DNA integrity and are the first line guardians of the genome against DNA breaks. Here we present a novel probe ([18F]-SuPAR) for noninvasive imaging of PARP-1/2 activity using positron emission tomography (PET). [18F]-SuPAR is a radiofluorinated nicotinamide adenine dinucleotide (NAD) analog that can be recognized by PARP-1/2 and incorporated into the long branched polymers of poly(ADP ribose) (PAR). The measurement of PARP-1/2 activity was supported by a reduction of radiotracer uptake in vivo following PARP-1/2 inhibition with talazoparib treatment, a potent PARP inhibitor recently approved by FDA for treatment of breast cancer, as well as ex vivo colocalization of radiotracer analog and poly(ADP ribose). With [18F]-SuPAR, we were able to map the dose- and time-dependent activation of PARP-1/2 following radiation therapy in breast and cervical cancer xenograft mouse models. Tumor response to therapy was determined by [18F]-SuPAR PET within 8 h of administration of a single dose of radiation equivalent to one round of stereotactic ablative radiotherapy.

  View details for DOI 10.1021/acs.bioconjchem.9b00089

  View details for Web of Science ID 000468368300008

  View details for PubMedID 30973715

• Engineered immune cells as highly sensitive cancer diagnostics NATURE BIOTECHNOLOGY Aalipour, A., Chuang, H., Murty, S., D'Souza, A. L., Park, S., Gulati, G. S., Patel, C. B., Beinat, C., Simonetta, F., Martinic, I., Gowrishankar, G., Robinson, E. R., Aalipour, E., Zhian, Z., Gambhir, S. S. 2019; 37 (5): 531-+

  View details for DOI 10.1038/s41587-019-0064-8

  View details for Web of Science ID 000469110000014

• A Non -Invasive Photoacoustic Imaging th Erythrocyte Derived Optical Nanoparticles to Detect CAD in In Vivo Mice Liu, Y., Hanley, T., Chen, H., Long, S., Gambhir, S., Cheng, Z., Wu, J., El Fakhri, G., Anvari, B., Zaman, R. SOC NUCLEAR MEDICINE INC. 2019

  View details for Web of Science ID 000473116800229

• Evaluation of Glycolytic Response to Multiple Classes of Anti-glioblastoma Drugs by Noninvasive Measurement of Pyruvate Kinase M2 Using [18F]DASA-23. Molecular imaging and biology : MIB : the official publication of the Academy of Molecular Imaging Beinat, C., Patel, C. B., Xie, Y., Gambhir, S. S. 2019

  Abstract

  PURPOSE: Pyruvate kinase M2 (PKM2) catalyzes the final step in glycolysis, the key process of tumor metabolism. PKM2 is found in high levels in glioblastoma (GBM) cells with marginal expression within healthy brain tissue, rendering it a key biomarker of GBM metabolic re-programming. Our group has reported the development of a novel radiotracer, 1-((2-fluoro- 6-[18F]fluorophenyl)sulfonyl)-4-((4-methoxyphenyl)sulfonyl)piperazine ([18F]DASA- 23), to non-invasively detect PKM2 levels with positron emission tomography (PET).PROCEDURE: U87 human GBM cells were treated with the IC50 concentration of various agents used in the treatment of GBM, including alkylating agents (temozolomide, carmustine, lomustine, procarbazine), inhibitor of topoisomerase I (irinotecan), vascular endothelial and epidermal growth factor receptor inhibitors (cediranib and erlotinib, respectively) anti-metabolite (5-fluorouracil), microtubule inhibitor (vincristine), and metabolic agents (dichloroacetate and IDH1 inhibitor ivosidenib). Following drug exposure for three or 6days (n=6 replicates per condition), the radiotracer uptake of [18F]DASA-23 and 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) was assessed. Changes in PKM2 protein levels were determined via Western blot and correlated to radiotracer uptake.RESULTS: Significant interactions were found between the treatment agent (n=12 conditions total comprised 11 drugs and vehicle) and the duration of treatment (3- or 6-day exposure to each drug) on the cellular uptake of [18F]DASA-23 (p=0.0001). The greatest change in the cellular uptake of [18F]DASA-23 was found after exposure to alkylating agents (p<0. 0001) followed by irinotecan (p=0. 0012), erlotinib (p=0. 02), and 5-fluorouracil (p=0. 005). Correlation of PKM2 protein levels and [18F]DASA-23 cellular uptake revealed a moderate correlation (r=0.44, p=0.15).CONCLUSIONS: These proof of principle studies emphasize the superiority of [18F]DASA-23 to [18F]FDG in detecting the glycolytic response of GBM to multiple classes of anti-neoplastic drugs in cell culture. A clinical trial evaluating the diagnostic utility of [18F]DASA-23 PET in GBM patients (NCT03539731) is ongoing.

  View details for PubMedID 30989436

• A Novel Engineered Small Protein for Positron Emission Tomography Imaging of Human Programmed Death Ligand-1: Validation in Mouse Models and Human Cancer Tissues. Clinical cancer research : an official journal of the American Association for Cancer Research Natarajan, A., Patel, C. B., Ramakrishnan, S., Panesar, P. S., Long, S. R., Gambhir, S. S. 2019; 25 (6): 1774-1785

  Abstract

  To design and evaluate a small engineered protein binder targeting human programmed death-1 ligand (hPD-L1) in vivo for PET imaging in four mouse tumor models, and in situ in human cancer specimens.Experimental Design: The hPD-L1 protein binder, FN3hPD-L1, was engineered using a 12-kDa human fibronectin type-3 domain (FN3) scaffold. The binder's affinity was assayed in CT26 mouse colon carcinoma cells stably expressing hPD-L1 (CT26/hPD-L1). 64Cu-FN3hPD-L1 was assayed for purity, specific activity, and immunoreactivity. Four groups of NSG mice (n = 3-5/group) were imaged with 64Cu-FN3hPD-L1 PET imaging (1-24 hours postinjection of 3.7 MBq/7 μg of Do-FN3 in 200 μL PBS): Nod SCID Gamma (NSG) mice bearing (i) syngeneic CT26/hPD-L1tumors, (ii) CT26/hPD-L1 tumors blocked (blk) by preinjected nonradioactive FN3hPD-L1 binder, (iii) hPD-L1-negative Raji xenografts, and (iv) MDA-MB-231 xenografts. The FN3hPD-L1 binder staining was evaluated against validated hPD-L1 antibodies by immunostaining in human cancer specimens.FN3hPD-L1 bound hPD-L1 with 1.4 ± 0.3 nmol/L affinity in CT26/hPD-L1 cells. 64Cu-FN3hPD-L1 radiotracer showed >70% yield and >95% purity. 64Cu-FN3hPD-L1 PET imaging of mice bearing CT26/hPD-L1 tumors showed tumor-to-muscle ratios of 5.6 ± 0.9 and 13.1 ± 2.3 at 1 and 4 hours postinjection, respectively. The FN3hPD-L1 binder detected hPD-L1 expression in human tissues with known hPD-L1 expression status based on two validated antibodies.The 64Cu-FN3hPD-L1 radiotracer represents a novel, small, and high-affinity binder for imaging hPD-L1 in tumors. Our data support further exploration and clinical translation of this binder for noninvasive identification of cancer patients who may respond to immune checkpoint blockade therapies.

  View details for DOI 10.1158/1078-0432.CCR-18-1871

  View details for PubMedID 30373750

  View details for PubMedCentralID PMC6420852

• Miniature gold nanorods for photoacoustic molecular imaging in the second near-infrared optical window. Nature nanotechnology Chen, Y., Zhao, Y., Yoon, S. J., Gambhir, S. S., Emelianov, S. 2019

  Abstract

  In photoacoustic imaging, the second near-infrared (NIR-II) window is where tissue generates the least background signal. However, the large size of the few available contrast agents in this spectral range impedes their pharmacokinetics and decreases their thermal stability, leading to unreliable photoacoustic imaging. Here, we report the synthesis of miniaturized gold nanorods absorbing in the NIR-II that are 5-11 times smaller than regular-sized gold nanorods with a similar aspect ratio. Under nanosecond pulsed laser illumination, small nanorods are about 3times more thermally stable and generate 3.5times stronger photoacoustic signal than their absorption-matched larger counterparts. These unexpected findings are confirmed using theoretical and numerical analysis, showing that photoacoustic signal is not only proportional to the optical absorption of the nanoparticle solution but also to the surface-to-volume ratio of the nanoparticles. In living tumour-bearing mice, these small targeted nanorods display a 30% improvement in efficiency of agent delivery to tumours and generate 4.5times greater photoacoustic contrast.

  View details for PubMedID 30833692

• Nanomedicine for Spontaneous Brain Tumors: A Companion Clinical Trial ACS NANO Arami, H., Patel, C. B., Madsen, S. J., Dickinson, P. J., Davis, R. M., Zeng, Y., Sturges, B. K., Woolard, K. D., Habte, F. G., Akin, D., Sinclair, R., Gambhir, S. S. 2019; 13 (3): 2858–69

  View details for DOI 10.1021/acsnano.8b04406

  View details for Web of Science ID 000462950500015

• Detection of Premalignant Gastrointestinal Lesions Using Surface-Enhanced Resonance Raman Scattering-Nanoparticle Endoscopy. ACS nano Harmsen, S., Rogalla, S., Huang, R., Spaliviero, M., Neuschmelting, V., Hayakawa, Y., Lee, Y., Tailor, Y., Toledo-Crow, R., Kang, J. W., Samii, J. M., Karabeber, H., Davis, R. M., White, J. R., van de Rijn, M., Gambhir, S. S., Contag, C. H., Wang, T. C., Kircher, M. F. 2019; 13 (2): 1354–64

  Abstract

  Cancers of the gastrointestinal (GI) tract are among the most frequent and most lethal cancers worldwide. An important reason for this high mortality is that early disease is typically asymptomatic, and patients often present with advanced, incurable disease. Even in high-risk patients who routinely undergo endoscopic screening, lesions can be missed due to their small size or subtle appearance. Thus, current imaging approaches lack the sensitivity and specificity to accurately detect incipient GI tract cancers. Here we report our finding that a single dose of a high-sensitivity surface-enhanced resonance Raman scattering nanoparticle (SERRS-NP) enables reliable detection of precancerous GI lesions in animal models that closely mimic disease development in humans. Some of these animal models have not been used previously to evaluate imaging probes for early cancer detection. The studies were performed using a commercial Raman imaging system, a newly developed mouse Raman endoscope, and finally a clinically applicable Raman endoscope for larger animal studies. We show that this SERRS-NP-based approach enables robust detection of small, premalignant lesions in animal models that faithfully recapitulate human esophageal, gastric, and colorectal tumorigenesis. This method holds promise for much earlier detection of GI cancers than currently possible and could lead therefore to marked reduction of morbidity and mortality of these tumor types.

  View details for PubMedID 30624916

• Assessment of Tumor Redox Status through (S)-4-(3-[F-18] fluoropropyl)-L-Glutamic Acid PET Imaging of System x(c)(-) Activity CANCER RESEARCH McCormick, P. N., Greenwood, H. E., Glaser, M., Maddocks, O. K., Gendron, T., Sander, K., Gowrishankar, G., Hoehne, A., Zhang, T., Shuhendler, A. J., Lewis, D. Y., Berndt, M., Koglin, N., Lythgoe, M. F., Gambhir, S. S., Arstad, E., Witney, T. H. 2019; 79 (4): 853–63

  View details for DOI 10.1158/0008-5472.CAN-18-2634

  View details for Web of Science ID 000458738900017

• In Vivo Translation of the CIRPI System---Revealing Molecular Pathology of Rabbit Aortic Atherosclerotic Plaques. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Zaman, R., Yousufi, S., Chibana, H., Ikeno, F., Long, S. R., Gambhir, S. S., Chin, F. T., McConnell, M. V., Xing, L., Yeung, A. 2019

  Abstract

  Introduction: Thin-cap fibro atheroma (TCFA), unstable lesions in coronary artery disease (CAD), that prones to rupture resulting in substantial morbidity and mortality worldwide. However, their small size and complex morphological/biological features make early detection and risk assessment difficult. To overcome this limitation, we tested our newly developed catheter-based Circumferential-Intravascular-Radioluminescence-Photoacoustic-Imaging (CIRPI) system in vivo rabbit abdominal aorta to detect and characterize TCFA. Methods: The CIRPI system includes a novel optical probe combining circumferential radioluminescence imaging (CRI) and photoacoustic tomography (PAT). The CIRPI system was tested in rabbit abdominal aorta in vivo (WHHL, n = 5) and controls (NZW, n = 2). Rabbits were fasted for 6 hours before 5.55*107 Bq 18F-FDG was injected one hour prior to the imaging procedure. The experiment was done under anesthetic. A bare metal stent was implanted in the dorsal abdominal aorta as landmark, followed by the 7F imaging catheters that were advanced up to the proximal stent edge (PSE). Our CIRPI and clinical OCT were performed using pullback and non-occlusive flushing techniques. Results were verified with histochemical analysis. Results: Our CIRPI system successfully detected the locations and characterized both stable and vulnerable aortic plaques in vivo among all WHHL rabbits. Calcification was detected from the stable plaque (540/560 nm), whereas TCFA exhibited phospholipids/cholesterol (1040 nm, 1210 nm). These findings were verified with clinical OCT showing an area of low attenuation filled with lipids within TCFA. PAT image illustrated broken elastic fiber/collagen that could be verified with the histochemical analysis. All WHHL rabbits exhibited sparse to severe macrophages. However, 4 WHHL rabbits showed both moderate to severe level of calcifications and cholesterol clefts. However, all rabbits exhibited broken elastic fibers and collagen deposition. Control rabbits showed normal wall thickness with no presence of plaque tissue compositions. These findings were verified with the OCT and histochemical analysis. Conclusion: Our novel multi-modality hybrid system has been successfully translated to in vivo evaluation of atherosclerotic plaque structure and biology in a pre-clinical rabbit models. This proposed a paradigm shift that unites molecular and pathologic imaging technologies. Therefore, it may enhance the clinical evaluation of TCFA, as well as expand our understanding of CAD.

  View details for PubMedID 30737298

• Nanomedicine for Spontaneous Brain Tumors: A Companion Clinical Trial. ACS nano Arami, H., Patel, C. B., Madsen, S. J., Dickinson, P. J., Davis, R. M., Zeng, Y., Sturges, B. K., Woolard, K. D., Habte, F. G., Akin, D., Sinclair, R., Gambhir, S. S. 2019

  Abstract

  Nanoparticles' enhanced permeation and retention (EPR) variations due to tumor heterogeneity in naturally occurring brain tumors are commonly neglected in preclinical nanomedicine studies. Recent pathological studies have shown striking similarities between brain tumors in humans and dogs, indicating that canine brain tumors may be a valuable model to evaluate nanoparticles' EPR in this context. We recruited canine clinical cases with spontaneous brain tumors to investigate nanoparticles' EPR in different brain tumor pathologies using surface-enhanced Raman spectroscopy (SERS). We used gold nanoparticles due to their surface plasmon effect that enables their sensitive and microscopic resolution detection using the SERS technique. Raman microscopy of the resected tumors showed heterogeneous EPR of nanoparticles into oligodendrogliomas and meningiomas of different grades, without any detectable traces in necrotic parts of the tumors or normal brain. Raman observations were confirmed by scanning electron microscopy (SEM) and X-ray elemental analyses, which enabled localization of individual nanoparticles embedded in tumor tissues. Our results demonstrate nanoparticles' EPR and its variations in clinically relevant, spontaneous brain tumors. Such heterogeneities should be considered alongside routine preoperative imaging and histopathological analyses in order to accelerate clinical management of brain tumors using nanomedicine approaches.

  View details for PubMedID 30714717

• Detection of visually occult metastatic lymph nodes using molecularly targeted fluorescent imaging during surgical resection of pancreatic cancer. HPB : the official journal of the International Hepato Pancreato Biliary Association Tummers, W. S., Miller, S. E., Teraphongphom, N. T., van den Berg, N. S., Hasan, A., Longacre, T. A., Fisher, G. A., Bonsing, B. A., Vahrmeijer, A. L., Gambhir, S. S., Swijnenburg, R., Rosenthal, E. L., Poultsides, G. A. 2019

  Abstract

  BACKGROUND: Although most patients with PDAC experience distant failure after resection, a significant portion still present with local recurrence. Intraoperative fluorescent imaging can potentially facilitate the visualization of involved peritumoral LNs and guide the locoregional extent of nodal dissection. Here, the efficacy of targeted intraoperative fluorescent imaging was examined in the detection of metastatic lymph nodes (LNs) during resection of pancreatic ductal adenocarcinoma (PDAC).METHODS: A dose-escalation prospective study was performed to assess feasibility of tumor detection within peripancreatic LNs using cetuximab-IRDye800 in PDAC patients. Fluorescent imaging of dissected LNs was analyzed exvivo macroscopically and microscopically and fluorescence was correlated with histopathology.RESULTS: A total of 144 LNs (72 in the low-dose and 72 in the high-dose cohort) were evaluated. Detection of metastatic LNs by fluorescence was better in the low-dose (50mg) cohort, where sensitivity and specificity was 100% and 78% macroscopically, and 91% and 66% microscopically. More importantly, this method was able to detect occult foci of tumor (measuring<5mm) with a sensitivity of 88% (15/17 LNs).CONCLUSION: This study provides proof of concept that intraoperative fluorescent imaging with cetuximab-IRDye800 can facilitate the detection of peripancreatic lymph nodes often containing subclinical foci of disease.

  View details for PubMedID 30723062

• Detection of Premalignant Gastrointestinal Lesions Using Surface-Enhanced Resonance Raman Scattering-Nanoparticle Endoscopy ACS NANO Harmsen, S., Rogalla, S., Huang, R., Spaliviero, M., Neuschmelting, V., Hayakawa, Y., Lee, Y., Tailor, Y., Toledo-Crow, R., Kang, J., Samii, J. M., Karabeber, H., Davis, R. M., White, J. R., van de Rijn, M., Gambhir, S. S., Contag, C. H., Wang, T. C., Kircher, M. F. 2019; 13 (2): 1354–64

  View details for DOI 10.1021/acsnano.8b06808

  View details for Web of Science ID 000460199400038

• First-in-human liver-tumour surgery guided by multispectral fluorescence imaging in the visible and near-infrared-I/II windows. Nature biomedical engineering Hu, Z. n., Fang, C. n., Li, B. n., Zhang, Z. n., Cao, C. n., Cai, M. n., Su, S. n., Sun, X. n., Shi, X. n., Li, C. n., Zhou, T. n., Zhang, Y. n., Chi, C. n., He, P. n., Xia, X. n., Chen, Y. n., Gambhir, S. S., Cheng, Z. n., Tian, J. n. 2019

  Abstract

  The second near-infrared wavelength window (NIR-II, 1,000-1,700 nm) enables fluorescence imaging of tissue with enhanced contrast at depths of millimetres and at micrometre-scale resolution. However, the lack of clinically viable NIR-II equipment has hindered the clinical translation of NIR-II imaging. Here, we describe an optical-imaging instrument that integrates a visible multispectral imaging system with the detection of NIR-II and NIR-I (700-900 nm in wavelength) fluorescence (by using the dye indocyanine green) for aiding the fluorescence-guided surgical resection of primary and metastatic liver tumours in 23 patients. We found that, compared with NIR-I imaging, intraoperative NIR-II imaging provided a higher tumour-detection sensitivity (100% versus 90.6%; with 95% confidence intervals of 89.1%-100% and 75.0%-98.0%, respectively), a higher tumour-to-normal-liver-tissue signal ratio (5.33 versus 1.45) and an enhanced tumour-detection rate (56.41% versus 46.15%). We infer that combining the NIR-I/II spectral windows and suitable fluorescence probes might improve image-guided surgery in the clinic.

  View details for DOI 10.1038/s41551-019-0494-0

  View details for PubMedID 31873212

• Improved detection of prostate cancer using a magneto-nanosensor assay for serum circulating autoantibodies. PloS one Xu, L., Lee, J., Hao, S., Ling, X. B., Brooks, J. D., Wang, S. X., Gambhir, S. S. 2019; 14 (8): e0221051

  Abstract

  PURPOSE: To develop a magneto-nanosensor (MNS) based multiplex assay to measure protein and autoantibody biomarkers from human serum for prostate cancer (CaP) diagnosis.MATERIALS AND METHODS: A 4-panel MNS autoantibody assay and a MNS protein assay were developed and optimized in our labs. Using these assays, serum concentration of six biomarkers including prostate-specific antigen (PSA) protein, free/total PSA ratio, as well as four autoantibodies against Parkinson disease 7 (PARK7), TAR DNA-binding protein 43 (TARDBP), Talin 1 (TLN1), and Caldesmon 1 (CALD1) and were analyzed. Human serum samples from 99 patients (50 with non-cancer and 49 with clinically localized CaP) were evaluated.RESULTS: The MNS assay showed excellent performance characteristics and no cross-reactivity. All autoantibody assays showed a statistically significant difference between CaP and non-cancer samples except for PARK7. The most significant difference was the combination of the four autoantibodies as a panel in addition to the free/total PSA ratio. This combination had the highest area under the curve (AUC)- 0.916 in ROC analysis.CONCLUSIONS: Our results suggest that this autoantibody panel along with PSA and free PSA have potential to segregate patients without cancer from those with prostate cancer with higher sensitivity and specificity than PSA alone.

  View details for DOI 10.1371/journal.pone.0221051

  View details for PubMedID 31404106

• Engineered immune cells as highly sensitive cancer diagnostics. Nature biotechnology Aalipour, A. n., Chuang, H. Y., Murty, S. n., D'Souza, A. L., Park, S. M., Gulati, G. S., Patel, C. B., Beinat, C. n., Simonetta, F. n., Martinić, I. n., Gowrishankar, G. n., Robinson, E. R., Aalipour, E. n., Zhian, Z. n., Gambhir, S. S. 2019

  Abstract

  Endogenous biomarkers remain at the forefront of early disease detection efforts, but many lack the sensitivities and specificities necessary to influence disease management. Here, we describe a cell-based in vivo sensor for highly sensitive early cancer detection. We engineer macrophages to produce a synthetic reporter on adopting an M2 tumor-associated metabolic profile by coupling luciferase expression to activation of the arginase-1 promoter. After adoptive transfer in colorectal and breast mouse tumor models, the engineered macrophages migrated to the tumors and activated arginase-1 so that they could be detected by bioluminescence imaging and luciferase measured in the blood. The macrophage sensor detected tumors as small as 25-50 mm3 by blood luciferase measurements, even in the presence of concomitant inflammation, and was more sensitive than clinically used protein and nucleic acid cancer biomarkers. Macrophage sensors also effectively tracked the immunological response in muscle and lung models of inflammation, suggesting the potential utility of this approach in disease states other than cancer.

  View details for PubMedID 30886438

• Engineered Immune Cells as Highly Sensitive Cancer Diagnostics Nature Biotechnology Aalipour, A., Chuang, H., Murty, S., D'Souza, A. L., Park, S., Gulanti, G. S., Patel, C. B., Beinat, C., Simonetta, F., Martinie, I., Gowrishankar, G., Robinson, E. R., Aalipour, E., Zhian, Z., Gambhir, S. S. 2019
• The characterization of 18F-hGTS13 for molecular imaging of xC- transporter activity with positron emission tomography. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Beinat, C. n., Gowrishankar, G. n., Shen, B. n., Alam, I. S., Robinson, E. n., Haywood, T. n., Patel, C. B., Azevedo, E. C., Castillo, J. n., Ilovich, O. n., Koglin, N. n., Schmitt-Willich, H. n., Berndt, M. n., Mueller, A. n., Zerna, M. n., Srinivasan, A. n., Gambhir, S. S. 2019

  Abstract

  Purpose: The aim of this study was development of an improved positron emission tomography (PET) radiotracer for measuring xC- activity with increased tumor uptake and reduced uptake in inflammatory cells compared to (S)-4-(3-18F-Fluoropropyl)-L-glutamic acid (18F-FSPG). Experimental design: A racemic glutamate derivative, 18F-hGTS13 was evaluated in cell culture and animal tumor models. 18F-hGTS13 was separated into C5-epimers and the corresponding 18F-hGTS13-isomer1 and 18F-hGTS13-isomer2 evaluated in H460 tumor bearing rats. Preliminary studies investigate the cellular uptake of 18F-hGTS13-isomer2 in multiple immune cell populations and states. Results:18F-hGTS13 demonstrated excellent H460 tumor visualization with high tumor-to-background ratios, confirmed by ex vivo biodistribution studies. Tumor associated radioactivity of 18F-hGTS13 (7.5±0.9%ID/g, n = 3) was significantly higher than with 18F-FSPG (4.6±0.7%ID/g, n = 3, P = 0.01). 18F-hGTS13-isomer2 exhibited excellent H460 tumor visualization (6.3±1.1%ID/g, n-3), and significantly reduced uptake in multiple immune cell populations relative to 18F-FSPG. 18F-hGTS13-isomer2 exhibited increased liver uptake relative to 18F-FSPG (4.6±0.8%ID/g vs. 0.7±0.01%ID/g) limiting its application in hepatocellular carcinoma. Conclusion:18F-hGTS13-isomer2 is a new PET radiotracer for molecular imaging of xC- activity which may provide information regarding tumor oxidation states. 18F-hGTS13-isomer2 has potential for clinical translation for imaging cancers of the thorax due to the low background signal in healthy tissue.

  View details for DOI 10.2967/jnumed.119.225870

  View details for PubMedID 31171595

• Initial experience with a PET/computed tomography system using silicon photomultiplier detectors. Nuclear medicine communications Park, S. Y., Barrato, L. n., Hatami, N. n., Davidzon, G. n., Gambhir, S. S., Iagaru, A. n. 2019

  Abstract

  A PET/computed tomography (CT) that uses silicon photomultiplier (SiPM) technology was installed at our institution. Here, we report the initial use of the new scanner and evaluate the image quality in comparison to standard PET/CT scanners.Seventy-two patients were scanned first using standard PET/CT followed immediately by the new PET/CT system. Images from the new PET/CT system were reconstructed using a conventional [non time-of-flight (TOF)] algorithm, TOF alone and TOF in combination with BSREM. Images from standard PET/CT were reconstructed using clinical standard-of-care settings. Three blinded readers randomly reviewed four datasets (standard, non-TOF, TOF alone, TOF+BSREM) per patient for image quality using a five-point Likert scale. SUV measurements for the single most avid lesion on each dataset were also recorded.Datasets from the new scanner had higher image quality (P < 0.001) and SUV measurements (P < 0.001) compared with the standard scanners, and scores further improved when TOF and BSREM algorithms were added (mean scores for standard, non-TOF, TOF alone and TOF+BSREM were 3.1, 3.9, 4.3 and 5.0, respectively; mean SUVmax for hottest lesion were 8.8, 10.3, 10.7 and 13.3, respectively).The SiPM-based PET/CT system outperforms two standard Bismuth germanium oxide- and Lutetium-yttrium oxyorthosilicate-based scanners in terms of image quality, with further benefits added using TOF and BSREM. This may be beneficial for detecting small lesions and more accurate disease staging.

  View details for DOI 10.1097/MNM.0000000000001088

  View details for PubMedID 31568189

• Positron emission tomography reporter gene strategy for use in the central nervous system PNAS Haywood, T., Beinat, C., Gowrishankar, G., Patel, C. B., Alam, I. S., Murty, S., Gambhir, S. S. 2019

  View details for DOI 10.1073/pnas.1901645116

• How to Prevent a Leaky Pipeline in Academic Radiology: Insights From a Faculty Survey. Journal of the American College of Radiology : JACR Daldrup-Link, H. n., Villavasso, K. n., Zhao, Q. n., Lu, Y. n., Ranieri, A. n., Simard, C. n., Gambhir, S. S. 2019

  View details for PubMedID 31092345

• Discussions with Leaders: A Conversation Between Sam Gambhir and Johannes Czernin. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Czernin, J., Gambhir, S. S. 2018; 59 (12): 1783–85

  View details for PubMedID 30510073

• Discussions with Leaders: A Conversation Between Sam Gambhir and Johannes Czernin JOURNAL OF NUCLEAR MEDICINE Czernin, J., Gambhir, S. 2018; 59 (12): 1783–85

  View details for DOI 10.2967/jnumed.118.221648

  View details for Web of Science ID 000452015900005

• Striatal dopamine deficits predict reductions in striatal functional connectivity in major depression: a concurrent 11C-raclopride positron emission tomography and functional magnetic resonance imaging investigation. Translational psychiatry Hamilton, J. P., Sacchet, M. D., Hjornevik, T., Chin, F. T., Shen, B., Kampe, R., Park, J. H., Knutson, B. D., Williams, L. M., Borg, N., Zaharchuk, G., Camacho, M. C., Mackey, S., Heilig, M., Drevets, W. C., Glover, G. H., Gambhir, S. S., Gotlib, I. H. 2018; 8 (1): 264

  Abstract

  Major depressive disorder (MDD) is characterized by the altered integration of reward histories and reduced responding of the striatum. We have posited that this reduced striatal activation in MDD is due to tonically decreased stimulation of striatal dopamine synapses which results in decremented propagation of information along the cortico-striatal-pallido-thalamic (CSPT) spiral. In the present investigation, we tested predictions of this formulation by conducting concurrent functional magnetic resonance imaging (fMRI) and 11C-raclopride positron emission tomography (PET) in depressed and control (CTL) participants. We scanned 16 depressed and 14 CTL participants with simultaneous fMRI and 11C-raclopride PET. We estimated raclopride binding potential (BPND), voxel-wise, and compared MDD and CTL samples with respect to BPND in the striatum. Using striatal regions that showed significant between-group BPND differences as seeds, we conducted whole-brain functional connectivity analysis using the fMRI data and identified brain regions in each group in which connectivity with striatal seed regions scaled linearly with BPND from these regions. We observed increased BPND in the ventral striatum, bilaterally, and in the right dorsal striatum in the depressed participants. Further, we found that as BPND increased in both the left ventral striatum and right dorsal striatum in MDD, connectivity with the cortical targets of these regions (default-mode network and salience network, respectively) decreased. Deficits in stimulation of striatal dopamine receptors in MDD could account in part for the failure of transfer of information up the CSPT circuit in the pathophysiology of this disorder.

  View details for PubMedID 30504860

• Striatal dopamine deficits predict reductions in striatal functional connectivity in major depression: a concurrent C-11-raclopride positron emission tomography and functional magnetic resonance imaging investigation TRANSLATIONAL PSYCHIATRY Hamilton, J., Sacchet, M. D., Hjornevik, T., Chin, F. T., Shen, B., Kampe, R., Park, J., Knutson, B. D., Williams, L. M., Borg, N., Zaharchuk, G., Camacho, M., Mackey, S., Heilig, M., Drevets, W. C., Glover, G. H., Gambhir, S. S., Gotlib, I. H. 2018; 8

  View details for DOI 10.1038/s41398-018-0316-2

  View details for Web of Science ID 000452318900002

• Tracking T Cell Activation By OX40 Immuno-PET: A Novel Strategy for Imaging of Graft Versus Host Disease Simonetta, F., Alam, I. S., Mayer, A. T., Murty, S., Vermesh, O., Hirai, T., Nobashi, T., Lau, K., Gambhir, S. S., Negrin, R. S. AMER SOC HEMATOLOGY. 2018

  View details for DOI 10.1182/blood-2018-99-116483

  View details for Web of Science ID 000454842804170

• Assessment of tumor redox status through (S)-4-(3-[18F]fluoropropyl)-L-glutamic acid positron emission tomography imaging of system xc- activity. Cancer research McCormick, P. N., Greenwood, H. E., Glaser, M., Maddocks, O. D., Gendron, T., Sander, K., Gowrishankar, G., Hoehne, A., Zhang, T., Shuhendler, A. J., Lewis, D. Y., Berndt, M., Koglin, N., Lythgoe, M. F., Gambhir, S. S., Arstad, E., Witney, T. H. 2018

  Abstract

  The cell's endogenous antioxidant system is vital to maintenance of redox homeostasis. Despite its central role in normal and pathophysiology, no non-invasive tools exist to measure this system in patients. The cystine/glutamate antiporter system xc- maintains the balance between intracellular reactive oxygen species and antioxidant production through the provision of cystine, a key precursor in glutathione biosynthesis. Here we show that tumor cell retention of a system xc--specific positron emission tomography radiotracer, (S)-4-(3-[18F]fluoropropyl)-L-glutamic acid ([18F]FSPG), decreases in proportion to levels of oxidative stress following treatment with a range of redox-active compounds. The decrease in [18F]FSPG retention correlated with a depletion of intracellular cystine resulting from increased de novo glutathione biosynthesis, shown through [U-13C6, U-15N2]cystine isotopic tracing. In vivo, treatment with the chemotherapeutic doxorubicin decreased [18F]FSPG tumor uptake in a mouse model of ovarian cancer, coinciding with markers of oxidative stress but preceding tumor shrinkage and decreased glucose utilization. Having already been used in pilot clinical trials, [18F]FSPG PET could be rapidly translated to the clinic as an early redox indicator of tumor response to treatment.

  View details for PubMedID 30401715

• A NOVEL METABOLIC PET TRACER STRATEGY TO DETERMINE EARLY EFFECTS OF TUMOR TREATING FIELDS (TTFIELDS) Patel, C., Beinat, C., Xie, Y., Haywood, T., Murty, S., Chang, E., Gambhir, S. OXFORD UNIV PRESS INC. 2018: 32

  View details for Web of Science ID 000460646300124

• Tumor Cell-Derived Extracellular Vesicle-Coated Nanocarriers: An Efficient Theranostic Platform for the Cancer-Specific Delivery of Anti-miR-21 and Imaging Agents ACS NANO Bose, R. C., Kumar, S., Zeng, Y., Afjei, R., Robinson, E., Lau, K., Bermudez, A., Habte, F., Pitteri, S. J., Sinclair, R., Willmann, J. K., Massoud, T. F., Gambhir, S. S., Paulmurugan, R. 2018; 12 (11): 10817–32

  View details for DOI 10.1021/acsnano.8b02587

  View details for Web of Science ID 000451789200022

• COMPARISON OF THREE METABOLIC PET RADIOTRACERS IN GLIOBLASTOMA: CELL CULTURE AND ANIMAL STUDIES Beinat, C., Patel, C., Murty, S., Haywood, T., Park, J., Xie, Y., Gambhir, S. OXFORD UNIV PRESS INC. 2018: 34

  View details for Web of Science ID 000460646300129

• EVALUATION OF GLYCOLYTIC RESPONSE TO SEVEN CLASSES OF ANTI-GLIOBLASTOMA DRUGS BY NON-INVASIVE MEASUREMENT OF PYRUVATE KINASE M2 Beinat, C., Patel, C., Xie, Y., Gambhir, S. OXFORD UNIV PRESS INC. 2018: 33–34

  View details for Web of Science ID 000460646300128

• Surface-Enhanced Raman Scattering Nanoparticles for Multiplexed Imaging of Bladder Cancer Tissue Permeability and Molecular Phenotype ACS NANO Davis, R. M., Kiss, B., Trivedi, D. R., Metzner, T. J., Liao, J. C., Gambhir, S. S. 2018; 12 (10): 9669–79

  View details for DOI 10.1021/acsnano.8b03217

  View details for Web of Science ID 000448751800006

• Emerging Intraoperative Imaging Modalities to Improve Surgical Precision MOLECULAR IMAGING AND BIOLOGY Alam, I. S., Steinberg, I., Vermesh, O., van den Berg, N. S., Rosenthal, E. L., van Dam, G. M., Ntziachristos, V., Gambhir, S. S., Hernot, S., Rogalla, S. 2018; 20 (5): 705–15

  View details for DOI 10.1007/s11307-018-1227-6

  View details for Web of Science ID 000444754000003

• Surface-Enhanced Raman Scattering Nanoparticles for Multiplexed Imaging of Bladder Cancer Tissue Permeability and Molecular Phenotype. ACS nano Davis, R. M., Kiss, B., Trivedi, D. R., Metzner, T. J., Liao, J. C., Gambhir, S. S. 2018

  Abstract

  Bladder cancer has the highest recurrence rate of all cancers due in part to inadequate transurethral resection. Inadequate resection is caused by the inability of cystoscopes to detect invisible lesions during the resection procedure. To improve detection and resection of nonmuscle invasive bladder cancer, we quantified the ability of a surface-enhanced Raman nanoparticle and endoscope system to classify bladder tissue as normal or cancerous. Both antibody-based (active) and tissue permeability-based (passive) targeting mechanisms were evaluated by topically applying nanoparticles to ex vivo human bladder tissue samples. Multiplexed molecular imaging of CD47 and Carbonic Anhydrase 9 tumor proteins gave a receiver operating characteristic area under the curve (ROC AUC of 0.93 (0.75, 1.00). Furthermore, passively targeted nanoparticles enabled tissue classification with an ROC AUC of 0.93 (0.73, 1.00). Passively targeted nanoparticles penetrated 5-fold deeper and bound to tumor tissue at 3.3-fold higher concentrations in cancer compared to normal bladder urothelium, suggesting the existence of an enhanced surface permeability and retention effect in human bladder cancer.

  View details for PubMedID 30203645

• Development and MPI tracking of novel hypoxia-targeted theranostic exosomes. Biomaterials Jung, K. O., Jo, H., Yu, J. H., Gambhir, S. S., Pratx, G. 2018; 177: 139–48

  Abstract

  Treating the hypoxic region of the tumor remains a significant challenge. The goals of this study are to develop an exosome platform that can target regions of tumor hypoxia and that can be monitored invivo using magnetic particle imaging (MPI). Four types of exosomes (generated under hypoxic or normoxic conditions, and with or without exposure to X-ray radiation) were isolated from MDA-MB-231 human breast cancer cells. Exosomes were labeled by DiO, a fluorescent lipophilic tracer, to quantify their uptake by hypoxic cancer cells. Subsequently, the exosomes were modified to carry SPIO (superparamagnetic iron oxide) nanoparticles and Olaparib (PARP inhibitor). FACS and fluorescence microscopy showed that hypoxic cells preferentially take up exosomes released by hypoxic cells, compared with other exosome formulations. In addition, the distribution of SPIO-labeled exosomes was successively imaged invivo using MPI. Finally, the therapeutic efficacy of Olaparib-loaded exosomes was demonstrated by increased apoptosis and slower tumor growth invivo. Our novel theranostic platform could be used as an effective strategy to monitor exosomes invivo and deliver therapeutics to hypoxic tumors.

  View details for PubMedID 29890363

• An intravascular magnetic wire for the high-throughput retrieval of circulating tumour cells in vivo. Nature biomedical engineering Vermesh, O., Aalipour, A., Ge, T. J., Saenz, Y., Guo, Y., Alam, I. S., Park, S. M., Adelson, C. N., Mitsutake, Y., Vilches-Moure, J., Godoy, E., Bachmann, M. H., Ooi, C. C., Lyons, J. K., Mueller, K., Arami, H., Green, A., Solomon, E. I., Wang, S. X., Gambhir, S. S. 2018; 2 (9): 696-705

  Abstract

  The detection and analysis of rare blood biomarkers is necessary for early diagnosis of cancer and to facilitate the development of tailored therapies. However, current methods for the isolation of circulating tumour cells (CTCs) or nucleic acids present in a standard clinical sample of only 5-10 ml of blood provide inadequate yields for early cancer detection and comprehensive molecular profiling. Here, we report the development of a flexible magnetic wire that can retrieve rare biomarkers from the subject's blood in vivo at a much higher yield. The wire is inserted and removed through a standard intravenous catheter and captures biomarkers that have been previously labelled with injected magnetic particles. In a proof-of-concept experiment in a live porcine model, we demonstrate the in vivo labelling and single-pass capture of viable model CTCs in less than 10 s. The wire achieves capture efficiencies that correspond to enrichments of 10-80 times the amount of CTCs in a 5-ml blood draw, and 500-5,000 times the enrichments achieved using the commercially available Gilupi CellCollector.

  View details for DOI 10.1038/s41551-018-0257-3

  View details for PubMedID 30505627

  View details for PubMedCentralID PMC6261517

• Development and MPI tracking of novel hypoxia-targeted theranostic exosomes BIOMATERIALS Jung, K., Jo, H., Yu, J., Gambhir, S., Pratx, G. 2018; 177: 139–48

  View details for DOI 10.1016/j.biomaterials.2018.05.048

  View details for Web of Science ID 000439401700012

• An intravascular magnetic wire for the high-throughput retrieval of circulating tumour cells in vivo NATURE BIOMEDICAL ENGINEERING Vermesh, O., Aalipour, A., Ge, T., Saenz, Y., Guo, Y., Alam, I. S., Park, S., Adelson, C. N., Mitsutake, Y., Vilches-Moure, J., Godoy, E., Bachmann, M. H., Ooi, C., Lyons, J. K., Mueller, K., Arami, H., Green, A., Solomon, E., Wang, S. X., Gambhir, S. S. 2018; 2 (9): 696–705

  View details for DOI 10.1038/s41551-018-0257-3

  View details for Web of Science ID 000444282800014

• The Immunoimaging Toolbox JOURNAL OF NUCLEAR MEDICINE Mayer, A. T., Gambhir, S. S. 2018; 59 (8): 1174–82

  View details for DOI 10.2967/jnumed.116.185967

  View details for Web of Science ID 000440582000003

• Molecular imaging agents for ultrasound CURRENT OPINION IN CHEMICAL BIOLOGY Zlitni, A., Gambhir, S. S. 2018; 45: 113–20

  View details for DOI 10.1016/j.cbpa.2018.03.017

  View details for Web of Science ID 000441686700015

• Quantification of Cerenkov Luminescence Imaging (CLI) Comparable With 3-D PET Standard Measurements MOLECULAR IMAGING Habte, F., Natarajan, A., Paik, D. S., Gambhir, S. 2018; 17

  View details for DOI 10.1177/1536012118788637

  View details for Web of Science ID 000439959300001

• Intraoperative Pancreatic Cancer Detection using Tumor-Specific Multimodality Molecular Imaging. Annals of surgical oncology Tummers, W. S., Miller, S. E., Teraphongphom, N. T., Gomez, A., Steinberg, I., Huland, D. M., Hong, S., Kothapalli, S., Hasan, A., Ertsey, R., Bonsing, B. A., Vahrmeijer, A. L., Swijnenburg, R., Longacre, T. A., Fisher, G. A., Gambhir, S. S., Poultsides, G. A., Rosenthal, E. L. 2018; 25 (7): 1880–88

  Abstract

  BACKGROUND: Operative management of pancreatic ductal adenocarcinoma (PDAC) is complicated by several key decisions during the procedure. Identification of metastatic disease at the outset and, when none is found, complete (R0) resection of primary tumor are key to optimizing clinical outcomes. The use of tumor-targeted molecular imaging, based on photoacoustic and fluorescence optical imaging, can provide crucial information to the surgeon. The first-in-human use of multimodality molecular imaging for intraoperative detection of pancreatic cancer is reported using cetuximab-IRDye800, a near-infrared fluorescent agent that binds to epidermal growth factor receptor.METHODS: A dose-escalation study was performed to assess safety and feasibility of targeting and identifying PDAC in a tumor-specific manner using cetuximab-IRDye800 in patients undergoing surgical resection for pancreatic cancer. Patients received a loading dose of 100mg of unlabeled cetuximab before infusion of cetuximab-IRDye800 (50mg or 100mg). Multi-instrument fluorescence imaging was performed throughout the surgery in addition to fluorescence and photoacoustic imaging ex vivo.RESULTS: Seven patients with resectable pancreatic masses suspected to be PDAC were enrolled in this study. Fluorescence imaging successfully identified tumor with a significantly higher mean fluorescence intensity in the tumor (0.09±0.06) versus surrounding normal pancreatic tissue (0.02±0.01), and pancreatitis (0.04±0.01; p<0.001), with a sensitivity of 96.1% and specificity of 67.0%. The mean photoacoustic signal in the tumor site was 3.7-fold higher than surrounding tissue.CONCLUSIONS: The safety and feasibilty of intraoperative, tumor-specific detection of PDAC using cetuximab-IRDye800 with multimodal molecular imaging of the primary tumor and metastases was demonstrated.

  View details for PubMedID 29667116

• 6"-F-18-Fluoromaltotriose PET Evaluation in Escherichia Coli-Induced Myositis: Is There Uptake Saturation in Control? REPLY JOURNAL OF NUCLEAR MEDICINE Wardak, M., Gowrishankar, G., Gambhir, S. 2018; 59 (7): 1166–67

  View details for DOI 10.2967/jnumed.118.208736

  View details for Web of Science ID 000437237200042

• Intraoperative Pancreatic Cancer Detection using Tumor-Specific Multimodality Molecular Imaging ANNALS OF SURGICAL ONCOLOGY Tummers, W. S., Miller, S. E., Teraphongphom, N. T., Gomez, A., Steinberg, I., Huland, D. M., Hong, S., Kothapalli, S., Hasan, A., Ertsey, R., Bonsing, B. A., Vahrmeijer, A. L., Swijnenburg, R., Longacre, T. A., Fisher, G. A., Gambhir, S. S., Poultsides, G. A., Rosenthal, E. L. 2018; 25 (7): 1880–88

  View details for DOI 10.1245/s10434-018-6453-2

  View details for Web of Science ID 000433907300018

• A novel theranostic strategy for MMP-14 expressing glioblastomas impacts survival Mohanty, S., Chen, Z., Li, K., Morais, G., Klockow, J., Yerneni, K., Pisani, L., Chin, F., Mitra, S., Cheshier, S., Chang, E., Gambhir, S., Rao, J., Loadman, P. M., Falconer, R. A., Daldrup-Link, H. E. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-LB-004

  View details for Web of Science ID 000468818900177

• Positron emission tomography imaging of activated T cells by targeting OX40 reveals spatiotemporal immune dynamics and predicts response to in situ tumor vaccination Mayer, A. T., Alam, I. S., Sagiv-Barfi, I., Wang, K., Vermesh, O., Czerwinski, D. K., Johnson, E. M., James, M. L., Levy, R., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1538-7445.AM2018-3031

  View details for Web of Science ID 000468819500394

• Advances in Diagnostic and Intraoperative Molecular Imaging of Pancreatic Cancer PANCREAS Tummers, W. S., Willmann, J. K., Bonsing, B. A., Vahrmeijer, A. L., Gambhir, S. S., Swijnenburg, R. 2018; 47 (6): 675–89

  Abstract

  Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis. To improve outcomes, there is a critical need for improved tools for detection, accurate staging, and resectability assessment. This could improve patient stratification for the most optimal primary treatment modality. Molecular imaging, used in combination with tumor-specific imaging agents, can improve established imaging methods for PDAC. These novel, tumor-specific imaging agents developed to target specific biomarkers have the potential to specifically differentiate between malignant and benign diseases, such as pancreatitis. When these agents are coupled to various types of labels, this type of molecular imaging can provide integrated diagnostic, noninvasive imaging of PDAC as well as image-guided pancreatic surgery. This review provides a detailed overview of the current clinical imaging applications, upcoming molecular imaging strategies for PDAC, and potential targets for imaging, with an emphasis on intraoperative imaging applications.

  View details for DOI 10.1097/MPA.0000000000001075

  View details for Web of Science ID 000435963800011

  View details for PubMedID 29894417

  View details for PubMedCentralID PMC6003672

• Emerging Intraoperative Imaging Modalities to Improve Surgical Precision. Molecular imaging and biology : MIB : the official publication of the Academy of Molecular Imaging Alam, I. S., Steinberg, I., Vermesh, O., van den Berg, N. S., Rosenthal, E. L., van Dam, G. M., Ntziachristos, V., Gambhir, S. S., Hernot, S., Rogalla, S. 2018

  Abstract

  Intraoperative imaging (IOI) is performed to guide delineation and localization of regions of surgical interest. While oncological surgical planning predominantly utilizes x-ray computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound (US), intraoperative guidance mainly remains on surgeon interpretation and pathology for confirmation. Over the past decades however, intraoperative guidance has evolved significantly with the emergence of several novel imaging technologies, including fluorescence-, Raman, photoacoustic-, and radio-guided approaches. These modalities have demonstrated the potential to further optimize precision in surgical resection and improve clinical outcomes for patients. Not only can these technologies enhance our understanding of the disease, they can also yield large imaging datasets intraoperatively that can be analyzed by deep learning approaches for more rapid and accurate pathological diagnosis. Unfortunately, many of these novel technologies are still under preclinical or early clinical evaluation. Organizations like the Intra-Operative Imaging Study Group of the European Society for Molecular Imaging (ESMI) support interdisciplinary interactions with the aim to improve technical capabilities in the field, an approach that can succeed only if scientists, engineers, and physicians work closely together with industry and regulatory bodies to resolve roadblocks to clinical translation. In this review, we provide an overview of a variety of novel IOI technologies, discuss their challenges, and present future perspectives on the enormous potential of IOI for oncological surgical navigation.

  View details for PubMedID 29916118

• A Dual-Modality Hybrid Imaging System Harnesses Radioluminescence and Sound to Reveal Molecular Pathology of Atherosclerotic Plaques SCIENTIFIC REPORTS Zaman, R. T., Yousefi, S., Long, S. R., Saito, T., Mandella, M., Qiu, Z., Chen, R., Contag, C. H., Gambhir, S. S., Chin, F. T., Khuri-Yakub, B. T., McConnell, M. V., Shung, K., Xing, L. 2018; 8: 8992

  Abstract

  Atherosclerosis is a progressive inflammatory condition caused by an unstable lesion, called thin-cap fibro atheromata (TCFA) that underlies coronary artery disease (CAD)-one of the leading causes of death worldwide. Therefore, early clinical diagnosis and effective risk stratification is important for CAD management as well as preventing progression to catastrophic events. However, early detection could be difficult due to their small size, motion, obscuring 18F-FDG uptake by adjacent myocardium, and complex morphological/biological features. To overcome these limitations, we developed a catheter-based Circumferential-Intravascular-Radioluminescence-Photoacoustic-Imaging (CIRPI) system that can detect vulnerable plaques in coronary arteries and characterizes them with respect to pathology and biology. Our CIRPI system combined two imaging modalities: Circumferential Radioluminescence Imaging (CRI) and PhotoAcoustic Tomography (PAT) within a novel optical probe. The probe's CaF2:Eu based scintillating imaging window provides a 360° view of human (n = 7) and murine carotid (n = 10) arterial plaques by converting β-particles into visible photons during 18F-FDG decay. A 60× and 63× higher radioluminescent signals were detected from the human and murine plaque inflammations, respectively, compared to the control. The system's photoacoustic imaging provided a comprehensive analysis of the plaque compositions and its morphologic information. These results were further verified with IVIS-200, immunohistochemical analysis, and autoradiography.

  View details for PubMedID 29895966

• The Immuno-Imaging Toolbox. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Mayer, A. T., Gambhir, S. S. 2018

  Abstract

  The recent clinical success of cancer immunotherapy has renewed interest in the development of tools to image the immune system. In general, immunotherapies attempt to enable the body's own immune cells to seek out and destroy malignant disease. Molecular imaging of the cells and molecules which regulate immunity could provide unique insight into the mechanisms of action, and failure, of immunotherapies. In this review, we will collectively refer to the tools applied towards imaging the immune system as the immuno-imaging toolbox. The immuno-imaging toolbox is comprised of imaging hardware, software, and biological wetware which together enable dynamic and non-invasive visualization of immune response. Other recent reviews have focused on specific portions of the immuno-imaging toolbox, including advances in imaging hardware(1) and certain classes of imaging probes(2, 3). Here we will attempt to provide a comprehensive overview of the current state-of-the-art immuno-imaging toolbox with a focus on imaging strategies and their applications towards immunotherapy.

  View details for PubMedID 29794226

• A novel synthesis of 6 ''-[F-18]-fluoromaltotriose as a PET tracer for imaging bacterial infection JOURNAL OF LABELLED COMPOUNDS & RADIOPHARMACEUTICALS Namavari, M., Gowrishankar, G., Srinivasan, A., Gambhir, S. S., Haywood, T., Beinat, C. 2018; 61 (5): 408–14

  View details for DOI 10.1002/jlcr.3601

  View details for Web of Science ID 000431660100001

• Prospective Evaluation of Ga-68-RM2 PET/MRI in Patients with Biochemical Recurrence of Prostate Cancer and Negative Findings on Conventional Imaging JOURNAL OF NUCLEAR MEDICINE Minamimoto, R., Sonni, I., Hancock, S., Vasanawala, S., Loening, A., Gambhir, S. S., Iagaru, A. 2018; 59 (5): 803–8

  View details for DOI 10.2967/jnumed.117.197624

  View details for Web of Science ID 000431261700038

• The preclinical characterization of [F-18]hGTS13 for imaging of x(C)(-) transporter activity Beinat, C., Gowrishanker, G., Shen, B., Alam, I., Robinson, E., Ilovich, O., Koglin, N., Schmitt-Willich, H., Berndt, M., Mueller, A., Zerna, M., Srinivasan, A., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2018

  View details for Web of Science ID 000467489901109

• A Novel Positron Emission Tomography Reporter Gene/Reporter Probe for the Central Nervous System Haywood, T., Beinat, C., Gowrishankar, G., Patel, C., Alam, I., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2018

  View details for Web of Science ID 000467489900079

• SiPM-based vs LYSO-based Ga-68-DOTA-TATE PET/CT: Comparison of Semi-Quantitative Measurements in Normal Tissues and Lesions Baratto, L., Toriihara, A., Hatami, N., Davidzon, G., Srinivas, S., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2018

  View details for Web of Science ID 000467489900430

• PET imaging of OX40+activated T cells predicts therapeutic response in a murine cancer vaccine model Alam, I., Mayer, A., Sagiv-Barfi, I., Vermesh, O., Wang, K., Johnson, E., Czerwinski, D., James, M. L., Levy, R., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2018

  View details for Web of Science ID 000467489900074

• Molecular Imaging of Cardiovascular Infections with 6 ''-[F-18]- Fluoromaltotriose PET/CT Wardak, M., Gowrishankar, G., Zhao, X., Namavari, M., Liu, Y., Neofytou, E., Haywood, T., Wu, J., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2018

  View details for Web of Science ID 000467489900037

• Initial experience with a SiPM-based PET/CT scanner: influence of acquisition time on image quality EJNMMI PHYSICS Sonni, I., Baratto, L., Park, S., Hatami, N., Srinivas, S., Davidzon, G., Gambhir, S., Iagaru, A. 2018; 5: 9

  Abstract

  A newly introduced PET/CT scanner (Discovery Meaningful Insights-DMI, GE Healthcare) includes the silicon photomultiplier (SiPM) with time-of-flight (TOF) technology first used in the GE SIGNA PET/MRI. In this study, we investigated the impact of various acquisition times on image quality using this SiPM-based PET/CT.We reviewed data from 58 participants with cancer who were scanned using the DMI PET/CT scanner. The administered dosages ranged 295.3-429.9 MBq (mean ± SD 356.3 ± 37.4) and imaging started at 71-142 min (mean ± SD 101.41 ± 17.52) after administration of the radiopharmaceutical. The patients' BMI ranged 19.79-46.16 (mean ± SD 26.55 ± 5.53). We retrospectively reconstructed the raw TOF data at 30, 60, 90, and 120 s/bed and at the standard image acquisition time per clinical protocol (180 or 210 s/bed depending on BMI). Each reconstruction was reviewed blindly by two nuclear medicine physicians and scored 1-5 (1-poor, 5-excellent quality). The liver signal-to-noise ratio (SNR) was used as a quantitative measure of image quality.The average scores ± SD of the readers were 2.61 ± 0.83, 3.70 ± 0.92, 4.36 ± 0.82, 4.82 ± 0.39, and 4.91 ± 0.91 for the 30, 60, 90, and 120 s/bed and at standard acquisition time, respectively. Inter-reader agreement on image quality assessment was good, with a weighted kappa of 0.80 (95% CI 0.72-0.81). In the evaluation of the effects of time per bed acquisition on semi-quantitative measurements, we found that the only time point significantly different from the standard time were 30 and 60 s (both with P < 0.001). The effects of dose and BMI were not statistically significant (P = 0.195 and 0.098, respectively). There was a significant positive effect of time on SNR (P < 0.001), as well as a significant negative effect of weight (P < 0.001).Our results suggest that despite significant delays from injection to imaging (due to comparison with standard PET/CT) compared to standard clinical operations and even in a population with average BMI > 25, images can be acquired as fast as 90 s/bed using the SiPM PET/CT and still result in very good image quality (average score > 4).

  View details for PubMedID 29666972

• A blood biomarker for monitoring response to anti-EGFR therapy. Cancer biomarkers : section A of Disease markers Hughes, N. P., Xu, L., Nielsen, C. H., Chang, E., Hori, S. S., Natarajan, A., Lee, S., Kjar, A., Kani, K., Wang, S. X., Mallick, P., Gambhir, S. S. 2018

  Abstract

  BACKGROUND AND OBJECTIVE: To monitor therapies targeted to epidermal growth factor receptors (EGFR) in non-small cell lung cancer (NSCLC), we investigated Peroxiredoxin 6 (PRDX6) as a biomarker of response to anti-EGFR agents.METHODS: We studied cells that are sensitive (H3255, HCC827) or resistant (H1975, H460) to gefitinib. PRDX6 was examined with either gefitinib or vehicle treatment using enzyme-linked immunosorbent assays. We created xenograft models from one sensitive (HCC827) and one resistant cell line (H1975) and monitored serum PRDX6 levels during treatment.RESULTS: PRDX6 levels in cell media from sensitive cell lines increased significantly after gefitinib treatment vs. vehicle, whereas there was no significant difference for resistant lines. PRDX6 accumulation over time correlated positively with gefitinib sensitivity. Serum PRDX6 levels in gefitinib-sensitive xenograft models increased markedly during the first 24 hours of treatment and then decreased dramatically during the following 48 hours. Differences in serum PRDX6 levels between vehicle and gefitinib-treated animals could not be explained by differences in tumor burden.CONCLUSIONS: Our results show that changes in serum PRDX6 during the course of gefitinib treatment of xenograft models provide insight into tumor response and such an approach offers several advantages over imaging-based strategies for monitoring response to anti-EGFR agents.

  View details for PubMedID 29689709

• Smart-Dust-Nanorice for Enhancement of Endogenous Raman Signal, Contrast in Photoacoustic Imaging, and T2-Shortening in Magnetic Resonance Imaging. Small (Weinheim an der Bergstrasse, Germany) Pohling, C., Campbell, J. L., Larson, T. A., Van de Sompel, D., Levi, J., Bachmann, M. H., Bohndiek, S. E., Jokerst, J. V., Gambhir, S. S. 2018: e1703683

  Abstract

  Raman microspectroscopy provides chemo-selective image contrast, sub-micrometer resolution, and multiplexing capabilities. However, it suffers from weak signals resulting in image-acquisition times of up to several hours. Surface-enhanced Raman scattering (SERS) can dramatically enhance signals of molecules in close vicinity of metallic surfaces and overcome this limitation. Multimodal, SERS-active nanoparticles are usually labeled with Raman marker molecules, limiting SERS to the coating material. In order to realize multimodal imaging while acquiring the rich endogenous vibronic information of the specimen, a core-shell particle based on "Nanorice", where a spindle-shaped iron oxide core is encapsulated by a closed gold shell, is developed. An ultrathin layer of silica prevents agglomeration and unwanted chemical interaction with the specimen. This approach provides Raman signal enhancement due to plasmon resonance effects of the shell while the optical absorption in the near-infrared spectral region provides contrast in photoacoustic tomography. Finally, T2-relaxation of a magnetic resonance imaging (MRI) experiment is altered by taking advantage of the iron oxide core. The feasibility for Raman imaging is evaluated by nearfield simulations and experimental studies on the primate cell line COS1. MRI and photoacoustics are demonstrated in agarose phantoms illustrating the promising translational nature of this strategy for clinical applications in radiology.

  View details for DOI 10.1002/smll.201703683

  View details for PubMedID 29635739

• Molecular imaging agents for ultrasound. Current opinion in chemical biology Zlitni, A., Gambhir, S. S. 2018; 45: 113–20

  Abstract

  Ultrasound (US) imaging is a safe, sensitive and affordable imaging modality with a wide usage in the clinic. US signal can be further enhanced by using echogenic contrast agents (UCAs) which amplify the US signal. Developments in UCAs which are targeted to sites of disease allow the use of US imaging to provide molecular information. Unfortunately, traditional UCAs are too large to leave the vascular space limiting the application of molecular US to intravascular markers. In this mini review, we highlight the most recent reports on the application of molecular US imaging in the clinic and summarize the latest nanoparticle platforms used to develop nUCAs. We believe that the highlighted technologies will have a great impact on the evolution of the US imaging field.

  View details for PubMedID 29631121

• Reply: Optimizing Strategies for Immune Checkpoint Imaging with Immuno-PET in Preclinical Study. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Mayer, A. T., Gambhir, S. S. 2018; 59 (4): 711–12

  View details for PubMedID 29348318

• Development and Preclinical Validation of a Cysteine Knottin Peptide Targeting Integrin alpha v beta 6 for Near-infrared Fluorescent-guided Surgery in Pancreatic Cancer CLINICAL CANCER RESEARCH Tummers, W. S., Kimura, R. H., Abou-Elkacem, L., Beinat, C., Vahrmeijer, A. L., Swijnenburg, R., Willmann, J. K., Gambhir, S. S. 2018; 24 (7): 1667–76

  View details for DOI 10.1158/1078-0432.CCR-17-2491

  View details for Web of Science ID 000429050000017

• Optimizing Strategies for Immune Checkpoint Imaging with Immuno-PET in Preclinical Study REPLY JOURNAL OF NUCLEAR MEDICINE Mayer, A. T., Gambhir, S. S. 2018; 59 (4): 711–12

  View details for DOI 10.2967/jnumed.117.205153

  View details for Web of Science ID 000428981100042

• Thy1-Targeted Microbubbles for Ultrasound Molecular Imaging of Pancreatic Ductal Adenocarcinoma CLINICAL CANCER RESEARCH Abou-Elkacem, L., Wang, H., Chowdhury, S. M., Kimura, R. H., Bachawal, S. V., Gambhir, S. S., Tian, L., Willmann, J. K. 2018; 24 (7): 1574–85

  View details for DOI 10.1158/1078-0432.CCR-17-2057

  View details for Web of Science ID 000429050000009

• Tumor characterization by ultrasound-release of multiple protein and microRNA biomarkers, preclinical and clinical evidence PLOS ONE D'Souza, A. L., Chevillet, J. R., Ghanouni, P., Yan, X., Tewari, M., Gambhir, S. S. 2018; 13 (3): e0194268

  Abstract

  We have previously shown that low frequency ultrasound can release biomarkers from cells into the murine circulation enabling an amplification and localization of the released biomarker that could be used as a blood-based method to detect cancer earlier and monitor therapy. In this study, we further demonstrate that this technique could be used for characterization of tumors and/or identification of cellular masses of unknown origin due to the release of multiple protein and nucleic acid biomarkers in cells in culture, mice and patients. We sonicated colon (LS174T) and prostate (LNCaP) cancer cell lines in culture at a low frequency of 1 MHz and show that there were several-fold changes in multiple protein and microRNA (miRNA) abundance with treatment at various intensities and time. This release was dependent on the duration and intensity of the sonication for both cell lines. Significant increased release in biomarkers was also observed following tumor sonication in living mice bearing subcutaneous LS174T cell line xenografts (for proteins and nucleic acids) and in an experimental LS174T liver tumor model (for proteins only). Finally, we demonstrated this methodology of multiple biomarker release in patients undergoing ablation of uterine fibroids using MR guided high intensity focused ultrasound. Two protein biomarkers significantly increased in the plasma after the ultrasound treatment in 21 samples tested. This proof that ultrasound-amplification method works in soft tissue tumor models together with biomarker multiplexing, could allow for an effective non-invasive method for identification, characterization and localization of incidental lesions, cancer and other disease. Pre-treatment quantification of the biomarkers, allows for individualization of quantitative comparisons. This individualization of normal marker levels in this method allows for specificity of the biomarker-increase to each patient, tumor or organ being studied.

  View details for PubMedID 29547636

• A PET imaging approach for determining EGFR mutation status for improved lung cancer patient management SCIENCE TRANSLATIONAL MEDICINE Sun, X., Xiao, Z., Chen, G., Han, Z., Liu, Y., Zhang, C., Sun, Y., Song, Y., Wang, K., Fang, F., Wang, X., Lin, Y., Xu, L., Shao, L., Li, J., Cheng, Z., Gambhir, S., Shen, B. 2018; 10 (431)

  Abstract

  Tumor heterogeneity and changes in epidermal growth factor receptor (EGFR) mutation status over time challenge the design of effective EGFR tyrosine kinase inhibitor (TKI) treatment strategies for non-small cell lung cancer (NSCLC). Therefore, there is an urgent need to develop techniques for comprehensive tumor EGFR profiling in real time, particularly in lung cancer precision medicine trials. We report a positron emission tomography (PET) tracer, N-(3-chloro-4-fluorophenyl)-7-(2-(2-(2-(2-18F-fluoroethoxy) ethoxy) ethoxy) ethoxy)-6-methoxyquinazolin-4-amine (18F-MPG), with high specificity to activating EGFR mutant kinase. We evaluate the feasibility of using 18F-MPG PET for noninvasive imaging and quantification of EGFR-activating mutation status in preclinical models of NSCLC and in patients with primary and metastatic NSCLC tumors. 18F-MPG PET in NSCLC animal models showed a significant correlation (R2 = 0.9050) between 18F-MPG uptake and activating EGFR mutation status. In clinical studies with NSCLC patients (n = 75), the concordance between the detection of EGFR activation by 18F-MPG PET/computed tomography (CT) and tissue biopsy reached 84.29%. There was a greater response to EGFR-TKIs (81.58% versus 6.06%) and longer median progression-free survival (348 days versus 183 days) in NSCLC patients when 18F-MPG PET/CT SUVmax (maximum standard uptake value) was ≥2.23 versus <2.23. Our study demonstrates that 18F-MPG PET/CT is a powerful method for precise quantification of EGFR-activating mutation status in NSCLC patients, and it is a promising strategy for noninvasively identifying patients sensitive to EGFR-TKIs and for monitoring the efficacy of EGFR-TKI therapy.

  View details for PubMedID 29515002

• Toward achieving precision health SCIENCE TRANSLATIONAL MEDICINE Gambhir, S., Ge, T., Vermesh, O., Spitler, R. 2018; 10 (430)

  View details for DOI 10.1126/scitranslmed.aao3612

  View details for Web of Science ID 000426291800005

• Toward achieving precision health. Science translational medicine Gambhir, S. S., Ge, T. J., Vermesh, O., Spitler, R. 2018; 10 (430)

  Abstract

  Health care systems primarily focus on patients after they present with disease, not before. The emerging field of precision health encourages disease prevention and earlier detection by monitoring health and disease based on an individual's risk. Active participation in health care can be encouraged with continuous health-monitoring devices, providing a higher-resolution picture of human health and disease. However, the development of monitoring technologies must prioritize the collection of actionable data and long-term user engagement.

  View details for DOI 10.1126/scitranslmed.aao3612

  View details for PubMedID 29491186

  View details for PubMedCentralID PMC5985668

• [F-18] FSPG-PET reveals increased cystine/glutamate antiporter (xc-) activity in a mouse model of multiple sclerosis JOURNAL OF NEUROINFLAMMATION Hoehne, A., James, M. L., Alam, I. S., Ronald, J. A., Schneider, B., D'Souza, A., Witney, T. H., Andrews, L. E., Cropper, H. C., Behera, D., Gowrishankar, G., Ding, Z., Wyss-Coray, T., Chin, F. T., Biswal, S., Gambhir, S. S. 2018; 15

  View details for DOI 10.1186/s12974-018-1080-1

  View details for Web of Science ID 000425975200001

• [18F]FSPG-PET reveals increased cystine/glutamate antiporter (xc-) activity in a mouse model of multiple sclerosis. Journal of neuroinflammation Hoehne, A., James, M. L., Alam, I. S., Ronald, J. A., Schneider, B., D'Souza, A., Witney, T. H., Andrews, L. E., Cropper, H. C., Behera, D., Gowrishankar, G., Ding, Z., Wyss-Coray, T., Chin, F. T., Biswal, S., Gambhir, S. S. 2018; 15 (1): 55

  Abstract

  The cystine/glutamate antiporter (xc-) has been implicated in several neurological disorders and, specifically, in multiple sclerosis (MS) as a mediator of glutamate excitotoxicity and proinflammatory immune responses. We aimed to evaluate an xc-specific positron emission tomography (PET) radiotracer, (4S)-4-(3-[18F]fluoropropyl)-L-glutamate ([18F]FSPG), for its ability to allow non-invasive monitoring of xc- activity in a mouse model of MS.Experimental autoimmune encephalomyelitis (EAE) was induced in C57BL/6 mice by subcutaneous injection of myelin oligodendrocyte glycoprotein (MOG35-55) peptide in complete Freund's adjuvant (CFA) followed by pertussis toxin. Control mice received CFA emulsion and pertussis toxin without MOG peptide, while a separate cohort of naïve mice received no treatment. PET studies were performed to investigate the kinetics and distribution of [18F]FSPG in naïve, control, pre-symptomatic, and symptomatic EAE mice, compared to 18F-fluorodeoxyglucose ([18F]FDG). After final PET scans, each mouse was perfused and radioactivity in dissected tissues was measured using a gamma counter. Central nervous system (CNS) tissues were further analyzed using ex vivo autoradiography or western blot. [18F]FSPG uptake in human monocytes, and T cells pre- and post-activation was investigated in vitro.[18F]FSPG was found to be more sensitive than [18F]FDG at detecting pathological changes in the spinal cord and brain of EAE mice. Even before clinical signs of disease, a small but significant increase in [18F]FSPG signal was observed in the spinal cord of EAE mice compared to controls. This increase in PET signal became more pronounced in symptomatic EAE mice and was confirmed by ex vivo biodistribution and autoradiography. Likewise, in the brain of symptomatic EAE mice, [18F]FSPG uptake was significantly higher than controls, with the largest changes observed in the cerebellum. Western blot analyses of CNS tissues revealed a significant correlation between light chain of xc- (xCT) protein levels, the subunit of xc- credited with its transporter activity, and [18F]FSPG-PET signal. In vitro [18F]FSPG uptake studies suggest that both activated monocytes and T cells contribute to the observed in vivo PET signal.These data highlight the promise of [18F]FSPG-PET as a technique to provide insights into neuroimmune interactions in MS and the in vivo role of xc- in the development and progression of this disease, thus warranting further investigation.

  View details for DOI 10.1186/s12974-018-1080-1

  View details for PubMedID 29471880

  View details for PubMedCentralID PMC5822551

• Eradication of spontaneous malignancy by local immunotherapy. Science translational medicine Sagiv-Barfi, I., Czerwinski, D. K., Levy, S., Alam, I. S., Mayer, A. T., Gambhir, S. S., Levy, R. 2018; 10 (426)

  Abstract

  It has recently become apparent that the immune system can cure cancer. In some of these strategies, the antigen targets are preidentified and therapies are custom-made against these targets. In others, antibodies are used to remove the brakes of the immune system, allowing preexisting T cells to attack cancer cells. We have used another noncustomized approach called in situ vaccination. Immunoenhancing agents are injected locally into one site of tumor, thereby triggering a T cell immune response locally that then attacks cancer throughout the body. We have used a screening strategy in which the same syngeneic tumor is implanted at two separate sites in the body. One tumor is then injected with the test agents, and the resulting immune response is detected by the regression of the distant, untreated tumor. Using this assay, the combination of unmethylated CG-enriched oligodeoxynucleotide (CpG)-a Toll-like receptor 9 (TLR9) ligand-and anti-OX40 antibody provided the most impressive results. TLRs are components of the innate immune system that recognize molecular patterns on pathogens. Low doses of CpG injected into a tumor induce the expression of OX40 on CD4+ T cells in the microenvironment in mouse or human tumors. An agonistic anti-OX40 antibody can then trigger a T cell immune response, which is specific to the antigens of the injected tumor. Remarkably, this combination of a TLR ligand and an anti-OX40 antibody can cure multiple types of cancer and prevent spontaneous genetically driven cancers.

  View details for DOI 10.1126/scitranslmed.aan4488

  View details for PubMedID 29386357

  View details for PubMedCentralID PMC5997264

• Dosimetry Prediction for Clinical Translation of 64Cu-Pembrolizumab ImmunoPET Targeting Human PD-1 Expression. Scientific reports Natarajan, A., Patel, C. B., Habte, F., Gambhir, S. S. 2018; 8 (1): 633

  Abstract

  The immune checkpoint programmed death 1 receptor (PD-1) expressed on some tumor-infiltrating lymphocytes, and its ligand (PD-L1) expressed on tumor cells, enable cancers to evade the immune system. Blocking PD-1 with the monoclonal antibody pembrolizumab is a promising immunotherapy strategy. Thus, noninvasively quantifying the presence of PD-1 expression in the tumor microenvironment prior to initiation of immune checkpoint blockade may identify the patients likely to respond to therapy. We have developed a 64Cu-pembrolizumab radiotracer and evaluated human dosimetry. The tracer was utilized to image hPD-1 levels in two subcutaneous mouse models: (a) 293 T/hPD-1 cells xenografted into NOD-scid IL-2Rγnull mice (NSG/293 T/hPD-1) and (b) human peripheral blood mononuclear cells engrafted into NSG bearing A375 human melanoma tumors (hNSG/A375). In each mouse model two cohorts were evaluated (hPD-1 blockade with pembrolizumab [blk] and non-blocked [nblk]), for a total of four groups (n = 3-5/group). The xenograft-to-muscle ratio in the NSG/293 T/hPD-1 model at 24 h was significantly increased in the nblk group (7.0 ± 0.5) compared to the blk group (3.4 ± 0.9), p = 0.01. The radiotracer dosimetry evaluation (PET/CT ROI-based and ex vivo) in the hNSG/A375 model revealed the highest radiation burden to the liver. In summary, we validated the 64Cu-pembrolizumab tracer's specific hPD-1 receptor targeting and predicted human dosimetry.

  View details for DOI 10.1038/s41598-017-19123-x

  View details for PubMedID 29330552

• Thy1-Targeted Microbubbles for Ultrasound Molecular Imaging of Pancreatic Ductal Adenocarcinoma. Clinical cancer research : an official journal of the American Association for Cancer Research Abou-Elkacem, L. n., Wang, H. n., Chowdhury, S. M., Kimura, R. H., Bachawal, S. V., Gambhir, S. S., Tian, L. n., Willmann, J. K. 2018

  Abstract

  To engineer a dual human and murine Thy1-binding single-chain-antibody ligand (Thy1-scFv) for contrast microbubble-enhanced ultrasound molecular imaging of pancreatic ductal adenocarcinoma (PDAC). Thy1-scFv were engineered using yeast-surface-display techniques. Binding to soluble human and murine Thy1 and to Thy1-expressing cells was assessed by flow cytometry. Thy1-scFv was then attached to gas-filled microbubbles to create MB Thy1-scFv. Thy1 binding of MB Thy1-scFv to Thy1-expressing cells was evaluated under flow shear stress conditions in flow-chamber experiments. MB scFv-scrambled and MB Non-targeted were used as negative controls. All microbubble types were tested in both orthotopic human PDAC xenografts and transgenic PDAC mice in vivo. Results: Thy1-scFv had a K D of 3.4±0.36 nM for human and 9.2±1.7 nM for murine Thy1 and showed binding to both soluble and cellularly expressed Thy1. MB Thy1-scFv attached to Thy1 with high affinity compared to negative control microbubbles P<0.01) as assessed by flow cytometry. Similarly, flow-chamber studies showed significantly (P<0.01) higher binding of MB Thy1-scFv (3.0±0.81 MB/cell) to Thy1-expressing cells than MB scFv-scrambled (0.57±0.53) and MB Non-targeted (0.43±0.53). In vivo ultrasound molecular imaging using MB Thy1-scFv demonstrated significantly higher signal (P<0.01) in both orthotopic (5.32±1.59 a.u.) and transgenic PDAC (5.68±2.5 a.u.) mice compared to chronic pancreatitis (0.84±0.6 a.u.) and normal pancreas (0.67±0.71 a.u.). Ex vivo immunofluorescence confirmed significantly (P<0.01) increased Thy1 expression in PDAC compared to chronic pancreatitis and normal pancreas tissue. Conclusions: A dual human and murine Thy1-binding scFv was designed to generate contrast microbubbles to allow PDAC detection with ultrasound.

  View details for PubMedID 29301827

• The Utility of [18F]DASA-23 for Molecular Imaging of Prostate Cancer with Positron Emission Tomography Molecular Imaging and Biology Beinat, C., Haywood, T., Chen, Y., Patel, C. B., Alam, I. S., Murty, S., Gambhir, S. S. 2018; 20 (6)

  View details for DOI 10.1007/s11307-018-1194-y

• A blood biomarker for monitoring response to anti-EGFR therapy CANCER BIOMARKERS Hughes, N. P., Xu, L., Nielsen, C. H., Chang, E., Hori, S. S., Natarajan, A., Lee, S., Kjaer, A., Kani, K., Wang, S. X., Mallick, P., Gambhir, S. 2018; 22 (2): 333–44

  View details for DOI 10.3233/CBM-171149

  View details for Web of Science ID 000437251500016

• An intravascular magnetic wire for the high-throughput retrieval of circulating tumour cells in vivo. Nature biomedical engineering Vermesh, O., Aalipour, A., Ge, T. J., Saenz, Y., Guo, Y., Alam, I. S., Park, S., Adelson, C. N., Mitsutake, Y., Vilches-Moure, J., Godoy, E., Bachmann, M., Ooi, C. C., Lyons, J. K., Mueller, K., Arami, H., Green, A., Solomon, E. I., Wang, S. X., Gambhir, S. S. 2018; 2: 696–705

  Abstract

  The detection and analysis of rare blood biomarkers is necessary for early cancer diagnosis and to facilitate the development of tailored therapies. However, current methods for the isolation of circulating tumor cells (CTCs) or nucleic acids present in a standard clinical sample of only 5-10 mL of blood provide inadequate yields for early cancer detection and comprehensive molecular profiling. We have developed a flexible magnetic wire that can retrieve rare biomarkers from the subject's blood in vivo at a much higher yield. The wire is inserted and removed through a standard intravenous catheter and captures biomarkers that have been previously labeled with injected magnetic particles. In a proof-of-concept experiment in a live porcine model, we demonstrate the in vivo labeling and single-pass capture of viable model CTCs in less than 10 seconds. The wire achieves capture efficiencies that correspond to enrichments of 10-80 times the amount of CTCs in a 5-mL blood draw, and to 500-5,000 times the enrichments achieved by the commercially available Gilupi CellCollector.

  View details for PubMedID 30524876

• A Novel Engineered Small Protein for Positron Emission Tomography Imaging of Human Programmed Death Ligand-1 : Validation in Mouse Models and Human Cancer Tissues Clinical Cancer Res Natarajan, A., Patel, C. B., Ramakrishnan, S., Panesar, P. S., Long, S. R., Gambhir, S. S. 2018

  View details for DOI 10.1158/1078-0432.CCR-18-1871

• Tumor Cell-Derived Extracellular Vesicle-Coated Nanocarriers: An Efficient Theranostic Platform for the Cancer-Specific Delivery of Anti-miR-21 and Imaging Agents. ACS nano Jc Bose, R. n., Uday Kumar, S. n., Zeng, Y. n., Afjei, R. n., Robinson, E. n., Lau, K. n., Bermudez, A. n., Habte, F. n., Pitteri, S. J., Sinclair, R. n., Willmann, J. K., Massoud, T. F., Gambhir, S. S., Paulmurugan, R. n. 2018

  Abstract

  MicroRNAs are critical regulators of cancer initiation, progression, and dissemination. Extensive evidence suggests that the inhibition of over-expressed oncogenic miRNA function can be a robust strategy for anticancer therapy. However, in vivo targeted delivery of miRNA therapeutics to various types of cancers remains a major challenge. Inspired by their natural synthesis and cargo delivery capabilities, researchers have exploited tumor cell-derived extracellular vesicles (TEVs) for the cancer-targeted delivery of therapeutics and theranostics. Here, we investigate a TEV-based nanoplatform for multimodal miRNA delivery and phototherapy treatments as well as the magnetic resonance imaging of cancer. We demonstrated loading of anti-miR-21 that blocks the function of endogenous oncogenic miR-21 over-expressed in cancer cells into and subsequent delivery by TEVs derived from 4T1 cells. We also produced Cy5-anti-miR-21-loaded TEVs from two other cancer cell lines (HepG2 and SKBR3) and confirmed their robust homologous and heterologous transfection efficiency and intracellular Cy5-anti-miR-21 delivery. Additionally, TEV-mediated anti-miR-21 delivery attenuated doxorubicin (DOX) resistance in breast cancer cells with a 3-fold higher cell kill efficiency than in cells treated with DOX alone. We then investigated TEVs as a biomimetic source for the functionalization of gold-iron oxide nanoparticles (GIONs) and demonstrated nanotheranostic properties of TEV-GIONs in vitro. TEV-GIONs demonstrated excellent T2 contrast in in vitro magnetic resonance (MR) imaging and resulted in efficient photothermal effect in 4T1 cells. We also evaluated the biodistribution and theranostic property of anti-miR-21 loaded TEV-GIONs in vivo by labeling with indocyanine green near-infrared dye. We further validated the tumor specific accumulation of TEV-GIONs using MR imaging. Our findings demonstrate that the distribution pattern of the TEV-anti-miR-21-GIONs correlated well with the tumor-targeting capability as well as the activity and efficacy obtained in response to doxorubicin combination treatments. TEVs and TEV-GIONs are promising nanotheranostics for future applications in cancer molecular imaging and therapy.

  View details for PubMedID 30346694

• Tumor treating fields increases membrane permeability in glioblastoma cells. Cell death discovery Chang, E. n., Patel, C. B., Pohling, C. n., Young, C. n., Song, J. n., Flores, T. A., Zeng, Y. n., Joubert, L. M., Arami, H. n., Natarajan, A. n., Sinclair, R. n., Gambhir, S. S. 2018; 4: 113

  Abstract

  Glioblastoma is the most common yet most lethal of primary brain cancers with a one-year post-diagnosis survival rate of 65% and a five-year survival rate of barely 5%. Recently the U.S. Food and Drug Administration approved a novel fourth approach (in addition to surgery, radiation therapy, and chemotherapy) to treating glioblastoma; namely, tumor treating fields (TTFields). TTFields involves the delivery of alternating electric fields to the tumor but its mechanisms of action are not fully understood. Current theories involve TTFields disrupting mitosis due to interference with proper mitotic spindle assembly. We show that TTFields also alters cellular membrane structure thus rendering it more permeant to chemotherapeutics. Increased membrane permeability through the imposition of TTFields was shown by several approaches. For example, increased permeability was indicated through increased bioluminescence with TTFields exposure or with the increased binding and ingress of membrane-associating reagents such as Dextran-FITC or ethidium D or with the demonstration by scanning electron microscopy of augmented number and sizes of holes on the cellular membrane. Further investigations showed that increases in bioluminescence and membrane hole production with TTFields exposure disappeared by 24 h after cessation of alternating electric fields thus demonstrating that this phenomenom is reversible. Preliminary investigations showed that TTFields did not induce membrane holes in normal human fibroblasts thus suggesting that the phenomenom was specific to cancer cells. With TTFields, we present evidence showing augmented membrane accessibility by compounds such as 5-aminolevulinic acid, a reagent used intraoperatively to delineate tumor from normal tissue in glioblastoma patients. In addition, this mechanism helps to explain previous reports of additive and synergistic effects between TTFields and other chemotherapies. These findings have implications for the design of combination therapies in glioblastoma and other cancers and may significantly alter standard of care strategies for these diseases.

  View details for PubMedID 30534421

• The Immuno-Imaging Toolbox Journal of Nuclear Medicine Mayer, A. T., Gambhir, S. S. 2018; 59 (8): 1174-1182
• [18F]FSPG-PET Reveals Increased Cystine/Glutamate Antiporter (xc-) Activity in a Mouse Model of Multiple Sclerosis Journal of Neuroinflammation Hoehne, A., James, M. L., Alam, I. S., Ronald, J. A., Schneider, B., D'Souza, A., Witney, T. H., Andrews, L. E., Cropper, H. C., Behera, D., Gowrishankar, G., Ding, Z., Wyss-Coray, T., Chin, F. T., Biswal, S., Gambhir, S. S. 2018; 15 (1): 55
• Tumor Treating Fields Increases Membrane Permeability in Glioblastoma Cells Cell Death Discovery Chang, E., Patel, C. B., Pohling, C., Young, C., Song, J., Flores, T., Zeng, Y., Joubert, L. M., Arami, H., Natarajan, A., Sinclair, R., Gambhir, S. S. 2018; 4

  View details for DOI 10.1038/s41420-018-0130-x

• Immune Cell Therapy and Imaging of Glioblastoma Glioblastoma: State-of-the-Art Clinical Neuroimaging Murty, S., Gambhir, S. S. edited by Iv, M., Wintermark, M., Massoud, T. D. Nova Science Publishers. 2018
• Role of Imaging in Early-Phase Trials NOVEL DESIGNS OF EARLY PHASE TRIALS FOR CANCER THERAPEUTICS Greene, L., Srinivas, S., Park, S., Hatami, N., Nobashi, T., Baratto, L., Toriihara, A., Gambhir, S. S., Kummar, S., Takimoto, C. 2018: 129–49

  View details for DOI 10.1016/B978-0-12-812512-0.00010-5

  View details for Web of Science ID 000464871500011

• Ferumoxytol-based Dual-modality Imaging Probe for Detection of Stem Cell Transplant Rejection. Nanotheranostics Li, K. n., Chan, C. T., Nejadnik, H. n., Lenkov, O. D., Wolterman, C. n., Paulmurugan, R. n., Yang, H. n., Gambhir, S. S., Daldrup-Link, H. E. 2018; 2 (4): 306–19

  Abstract

  Purpose: Stem cell transplants are an effective approach to repair large bone defects. However, comprehensive techniques to monitor the fate of transplanted stem cells in vivo are lacking. Such strategies would enable corrective interventions at an early stage and greatly benefit the development of more successful tissue regeneration approaches. In this study, we designed and synthesized a dual-modality imaging probe (Feru-AFC) that can simultaneously localize transplanted stem cells and diagnose immune rejection-induced apoptosis at an early stage in vivo. Methods: We used a customized caspase-3 cleavable peptide-dye conjugate to modify the surface of clinically approved ferumoxytol nanoparticles (NPs) to generate the dual-modality imaging probe with fluorescence "light-up" feature. We labeled both mouse mesenchymal stem cells (mMSCs, matched) and pig mesenchymal stem cells (pMSCs, mismatched) with the probe and transplanted the labeled cells with biocompatible scaffold at the calvarial defects in mice. We then employed intravital microscopy (IVM) and magnetic resonance imaging (MRI) to investigate the localization, engraftment, and viability of matched and mismatched stem cells, followed by histological analyses to evaluate the results obtained from in vivo studies. Results: The Feru-AFC NPs showed good cellular uptake efficiency in the presence of lipofectin without cytotoxicity to mMSCs and pMSCs. The fluorescence of Feru-AFC NPs was turned on inside apoptotic cells due to the cleavage of peptide by activated caspase-3 and subsequent release of fluorescence dye molecules. Upon transplantation at the calvarial defects in mice, the intense fluorescence from the cleaved Feru-AFC NPs in apoptotic pMSCs was observed with a concomitant decrease in the overall cell number from days 1 to 6. In contrast, the Feru-AFC NP-treated mMSCs exhibited minimum fluorescence and the cell number also remained similar. Furthermore, in vivo MRI of the Feru-AFC NP-treated mMSC and pMSCs transplants could clearly indicate the localization of matched and mismatched cells, respectively. Conclusions: We successfully developed a dual-modality imaging probe for evaluation of the localization and viability of transplanted stem cells in mouse calvarial defects. Using ferumoxytol NPs as the platform, our Feru-AFC NPs are superparamagnetic and display a fluorescence "light-up" signature upon exposure to activated caspase-3. The results show that the probe is a promising tool for long-term stem cell tracking through MRI and early diagnosis of immune rejection-induced apoptosis through longitudinal fluorescence imaging.

  View details for DOI 10.7150/ntno.26389

  View details for PubMedID 29977742

  View details for PubMedCentralID PMC6030766

• Imaging activated T cells predicts response to cancer vaccines. The Journal of clinical investigation Alam, I. S., Mayer, A. T., Sagiv-Barfi, I. n., Wang, K. n., Vermesh, O. n., Czerwinski, D. K., Johnson, E. M., James, M. L., Levy, R. n., Gambhir, S. S. 2018

  Abstract

  In situ cancer vaccines are under active clinical investigation, given their reported ability to eradicate both local and disseminated malignancies. Intratumoral vaccine administration is thought to activate a T cell-mediated immune response, which begins in the treated tumor and cascades systemically. In this study, we describe a PET tracer (64Cu-DOTA-AbOX40) that enabled noninvasive and longitudinal imaging of OX40, a cell-surface marker of T cell activation. We report the spatiotemporal dynamics of T cell activation following in situ vaccination with CpG oligodeoxynucleotide in a dual tumor-bearing mouse model. We demonstrate that OX40 imaging was able to predict tumor responses on day 9 after treatment on the basis of tumor tracer uptake on day 2, with greater accuracy than both anatomical and blood-based measurements. These studies provide key insights into global T cell activation following local CpG treatment and indicate that 64Cu-DOTA-AbOX40 is a promising candidate for monitoring clinical cancer immunotherapy strategies.

  View details for PubMedID 29596062

• Quantification of Cerenkov Luminescence Imaging (CLI) Comparable With 3-D PET Standard Measurements. Molecular imaging Habte, F., Natarajan, A., Paik, D. S., Gambhir, S. S. 2018; 17: 1536012118788637

  Abstract

  Cerenkov luminescence imaging (CLI) is commonly performed using two-dimensional (2-D) conventional optical imaging systems for its cost-effective solution. However, quantification of CLI comparable to conventional three-dimensional positron emission tomography (PET) is challenging using these systems due to both the high attenuation of Cerenkov radiation (CR) on mouse tissue and nonexisting depth resolution of CLI using 2-D imaging systems (2-D CLI). In this study, we developed a model that estimates effective tissue attenuation coefficient and corrects the tissue attenuation of CLI signal intensity independent of tissue depth and size. To evaluate this model, we used several thin slices of ham as a phantom and placed a radionuclide (89Zr and 64Cu) inside the phantom at different tissue depths and sizes (2, 7, and 12 mm). We performed 2-D CLI and MicroPET/CT (Combined small animal PET and Computed Tomography (CT)) imaging of the phantom and in vivo mouse model after administration of 89Zr tracer. Estimates of the effective tissue attenuation coefficient (mueff) for 89Zr and 64Cu were 2.4 and 2.6 cm-1, respectively. The computed unit conversion factor to %ID/g from 2-D CLI signal was 2.74 * 10-3 muCi/radiance estimated from phantom study. After applying tissue attenuation correction and unit conversion to the in vivo animal study, an average quantification difference of 10% for spleen and 35% for liver was obtained compared to PET measurements. The proposed model provides comparable quantification accuracy to standard PET system independent of deep tissue CLI signal attenuation.

  View details for PubMedID 30043654

• Reply: 6"-18F-Fluoromaltotriose PET Evaluation in Escherichia-Coli-Induced Myositis: is there Uptake Saturation in Control? Journal of nuclear medicine : official publication, Society of Nuclear Medicine Wardak, M. n., Gowrishankar, G. n., Gambhir, S. S. 2018

  View details for PubMedID 29653976

• Intraoperative Molecular Imaging in Lung Cancer: The State of the Art and the Future. Molecular therapy : the journal of the American Society of Gene Therapy Rogalla, S. n., Joosten, S. C., Alam, I. S., Gambhir, S. S., Vermesh, O. n. 2018; 26 (2): 338–41

  View details for PubMedID 29398484

• The Utility of [18F]DASA-23 for Molecular Imaging of Prostate Cancer with Positron Emission Tomography. Molecular imaging and biology : MIB : the official publication of the Academy of Molecular Imaging Beinat, C. n., Haywood, T. n., Chen, Y. S., Patel, C. B., Alam, I. S., Murty, S. n., Gambhir, S. S. 2018

  Abstract

  There is a strong, unmet need for superior positron emission tomography (PET) imaging agents that are able to measure biochemical processes specific to prostate cancer. Pyruvate kinase M2 (PKM2) catalyzes the concluding step in glycolysis and is a key regulator of tumor growth and metabolism. Elevation of PKM2 expression was detected in Gleason 8-10 tumors compared to Gleason 6-7 carcinomas, indicating that PKM2 may potentially be a marker of aggressive prostate cancer. We have recently reported the development of a PKM2-specific radiopharmaceutical [18F]DASA-23 and herein describe its evaluation in cell culture and preclinical models of prostate cancer.The cellular uptake of [18F]DASA-23 was evaluated in a panel of prostate cancer cell lines and compared to that of [18F]FDG. The specificity of [18F]DASA-23 to measure PKM2 levels in cell culture was additionally confirmed through the use of PKM2-specific siRNA. PET imaging studies were then completed utilizing subcutaneous prostate cancer xenografts using either PC3 or DU145 cells in mice.[18F]DASA-23 uptake values over 60-min incubation period in PC3, LnCAP, and DU145 respectively were 23.4 ± 4.5, 18.0 ± 2.1, and 53.1 ± 4.6 % tracer/mg protein. Transient reduction in PKM2 protein expression with siRNA resulted in a 50.1 % reduction in radiotracer uptake in DU145 cells. Small animal PET imaging revealed 0.86 ± 0.13 and 1.6 ± 0.2 % ID/g at 30 min post injection of radioactivity in DU145 and PC3 subcutaneous tumor bearing mice respectively.Herein, we evaluated a F-18-labeled PKM2-specific radiotracer, [18F]DASA-23, for the molecular imaging of prostate cancer with PET. [18F]DASA-23 revealed rapid and extensive uptake levels in cellular uptake studies of prostate cancer cells; however, there was only modest tumor uptake when evaluated in mouse subcutaneous tumor models.

  View details for PubMedID 29736561

• Development and preclinical validation of a cysteine knottin peptide targeting Integrin αvβ6 for near-infrared fluorescent-guided surgery in pancreatic cancer. Clinical cancer research : an official journal of the American Association for Cancer Research Tummers, W. S., Kimura, R. H., Abou-Elkacem, L. n., Vahrmeijer, A. L., Swijnenburg, R. J., Willmann, J. K., Gambhir, S. S. 2018

  Abstract

  Intraoperative near-infrared fluorescence (NIRF) imaging could help stratification for the proper primary treatment for patients with pancreatic ductal adenocarcinoma (PDAC), and achieve complete resection since it allows visualization of cancer in real time. Integrin αvβ6, a target specific for PDAC, is present in >90% of patients, and is able to differentiate between pancreatitis and PDAC. A clinically translatable αvβ6-targeting NIRF agent was developed, based on a previously developed cysteine knottin peptide for PET imaging, R01-MG, and validated in preclinical mouse models.The applicability of the agent was tested for cell and tissue binding characteristics using cell-based plate assays, subcutaneous and orthotopic pancreatic models, and a transgenic mouse model of PDAC development (Pdx1-Cre tg/+;KRas LSL G12D/+;Ink4a/Arf). IRDye800CW was conjugated to R01-MG in a 1:1 ratio. R01-MG-IRDye800, was compared to a control peptide and IRDye800 alone.In subcutaneous tumor models a significantly higher tumor-to-background ratio (TBR) was seen in BxPC-3 tumors (2.5±0.1) compared to MiaPaCa-2 (1.2±0.1) (p<0.001), and to the control peptide (1.6±0.4) (p<0.005). In an orthotopic tumor model tumor-specific uptake of R01-MG-IRDye800 was shown compared to IRDye800 alone (TBR 2.7 versus 0.86). The fluorescent signal in tumors of transgenic mice was significantly higher, TBR of 3.6±0.94, compared to the normal pancreas of wild type controls, TBR of 1.0±0.17 (p<0.001).R01-MG-IRDye800 shows specific targeting to αvβ6, and holds promise as a diagnostic and therapeutic tool to recognize PDAC for fluorescence-guided surgery. This agent can help improve the stratification of patients for a potentially curative, margin-negative resection.

  View details for PubMedID 29298796

• A novel synthesis of 6''-[18 F]-fluoromaltotriose as a PET tracer for imaging bacterial infection. Journal of labelled compounds & radiopharmaceuticals Namavari, M. n., Gowrishankar, G. n., Srinivasan, A. n., Gambhir, S. S. 2018

  Abstract

  The aim of this study was to develop a positron emission tomography (PET) tracer to visualize and monitor therapeutic response to bacterial infections. In our continued efforts to find maltose based PET tracers that can image bacterial infections, we have designed and prepared 6''-[18 F]fluoromaltotriose as a second generation PET imaging tracer targeting the maltodextrin transporter of bacteria. We have developed methods to synthesize 6''-deoxy-6''-[18 F]fluoro-α-D-glucopyranosyl-(1-4)-O-α-D-glucopyranosyl-(1-4)-O-D-glucopyranose (6''-[18 F]-fluoromaltotriose) as a bacterial infection PET imaging agent. 6''-[18 F]fluoromaltotriose was prepared from precursor, 2'',3'',4''-tri-O-acetyl-6''-O-nosyl-α-D-glucopyranosyl-(1-4)-O-2',3',6'-tri-O-acetyl-α-D-glucopyranosyl-(1-4)-1,2,3,6-tetra-O-acetyl-D-glucopyranose (per-O-acetyl-6''-O-nosyl-maltotriose 4). This method utilizes the reaction between precursor 4 and anhydrous [18 F]KF/Kryptofix 2.2.2 in Dimethylformamide (DMF) at 85o C for 10 minutes to yield per-O-acetyl-6''-deoxy-6-'' [18 F]-fluoromaltotriose (7). Successive acidic and basic hydrolysis of the acetyl protecting groups in 7 produced 6''-[18 F]fluoromaltotriose (8). Also, cold 6''- [19 F]fluoromaltotriose was prepared from per-O-acetyl-6''-hydroxymaltotriose via a DAST reaction followed by a basic hydrolysis. A successful synthesis of 6''-[18 F]-fluoromaltotriose has been accomplished in 8±1.2 % radiochemical yield (decay corrected). Total synthesis time was 120 min. Serum stability of 6''-[18 F]fluoromaltotriose at 37o C indicated that 6''-[18 F]-fluoromaltotriose remained intact up to 2 h. In conclusion, we have successfully synthesized 6''-[18 F]-fluoromaltotriose via direct fluorination of an appropriate precursor of a protected maltotriose.

  View details for PubMedID 29314161

• Tomographic magnetic particle imaging of cancer targeted nanoparticles. Nanoscale Arami, H., Teeman, E., Troksa, A., Bradshaw, H., Saatchi, K., Tomitaka, A., Gambhir, S. S., Häfeli, U. O., Liggitt, D., Krishnan, K. M. 2017; 9 (47): 18723-18730

  Abstract

  Magnetic Particle Imaging (MPI) is an emerging, whole body biomedical imaging technique, with sub-millimeter spatial resolution and high sensitivity to a biocompatible contrast agent consisting of an iron oxide nanoparticle core and a biofunctionalized shell. Successful application of MPI for imaging of cancer depends on the nanoparticles (NPs) accumulating at tumors at sufficient levels relative to other sites. NPs' physiochemical properties such as size, crystallographic structure and uniformity, surface coating, stability, blood circulation time and magnetization determine the efficacy of their tumor accumulation and MPI signal generation. Here, we address these criteria by presenting strategies for the synthesis and surface functionalization of efficient MPI tracers, that can target a typical murine brain cancer model and generate three dimensional images of these tumors with very high signal-to-noise ratios (SNR). Our results showed high contrast agent sensitivities that enabled us to detect 1.1 ng of iron (SNR ∼ 3.9) and enhance the spatial resolution to about 600 μm. The biodistribution of these NPs was also studied using near-infrared fluorescence (NIRF) and single-photon emission computed tomography (SPECT) imaging. NPs were mainly accumulated in the liver and spleen and did not show any renal clearance. This first pre-clinical study of cancer targeted NPs imaged using a tomographic MPI system in an animal model paves the way to explore new nanomedicine strategies for cancer diagnosis and therapy, using clinically safe magnetic iron oxide nanoparticles and MPI.

  View details for DOI 10.1039/c7nr05502a

  View details for PubMedID 29165498

• SURFACE-ENHANCED RAMAN SPECTROSCOPY (SERS) FOR INTRAOPERATIVE BRAIN TUMOR IMAGING AND PHOTOTHERMAL THERAPY Arami, H., Chang, E., Patel, C. B., Madsen, S., Davis, R., Sinclair, R., Gambhir, S. S. OXFORD UNIV PRESS INC. 2017: 159

  View details for Web of Science ID 000415152503011

• Future cancer research priorities in the USA: a Lancet Oncology Commission. The Lancet. Oncology Jaffee, E. M., Dang, C. V., Agus, D. B., Alexander, B. M., Anderson, K. C., Ashworth, A., Barker, A. D., Bastani, R., Bhatia, S., Bluestone, J. A., Brawley, O., Butte, A. J., Coit, D. G., Davidson, N. E., Davis, M., DePinho, R. A., Diasio, R. B., Draetta, G., Frazier, A. L., Futreal, A., Gambhir, S. S., Ganz, P. A., Garraway, L., Gerson, S., Gupta, S., Heath, J., Hoffman, R. I., Hudis, C., Hughes-Halbert, C., Ibrahim, R., Jadvar, H., Kavanagh, B., Kittles, R., Le, Q. T., Lippman, S. M., Mankoff, D., Mardis, E. R., Mayer, D. K., McMasters, K., Meropol, N. J., Mitchell, B., Naredi, P., Ornish, D., Pawlik, T. M., Peppercorn, J., Pomper, M. G., Raghavan, D., Ritchie, C., Schwarz, S. W., Sullivan, R., Wahl, R., Wolchok, J. D., Wong, S. L., Yung, A. 2017; 18 (11): e653-e706

  Abstract

  We are in the midst of a technological revolution that is providing new insights into human biology and cancer. In this era of big data, we are amassing large amounts of information that is transforming how we approach cancer treatment and prevention. Enactment of the Cancer Moonshot within the 21st Century Cures Act in the USA arrived at a propitious moment in the advancement of knowledge, providing nearly US$2 billion of funding for cancer research and precision medicine. In 2016, the Blue Ribbon Panel (BRP) set out a roadmap of recommendations designed to exploit new advances in cancer diagnosis, prevention, and treatment. Those recommendations provided a high-level view of how to accelerate the conversion of new scientific discoveries into effective treatments and prevention for cancer. The US National Cancer Institute is already implementing some of those recommendations. As experts in the priority areas identified by the BRP, we bolster those recommendations to implement this important scientific roadmap. In this Commission, we examine the BRP recommendations in greater detail and expand the discussion to include additional priority areas, including surgical oncology, radiation oncology, imaging, health systems and health disparities, regulation and financing, population science, and oncopolicy. We prioritise areas of research in the USA that we believe would accelerate efforts to benefit patients with cancer. Finally, we hope the recommendations in this report will facilitate new international collaborations to further enhance global efforts in cancer control.

  View details for DOI 10.1016/S1470-2045(17)30698-8

  View details for PubMedID 29208398

  View details for PubMedCentralID PMC6178838

• A Pixel Pitch-Matched Ultrasound Receiver for 3-D Photoacoustic Imaging With Integrated Delta-Sigma Beamformer in 28-nm UTBB FD-SOI. IEEE journal of solid-state circuits Chen, M. C., Perez, A. P., Kothapalli, S. R., Cathelin, P., Cathelin, A., Gambhir, S. S., Murmann, B. 2017; 52 (11): 2843-2856

  Abstract

  This paper presents a pixel pitch-matched readout chip for 3-D photoacoustic (PA) imaging, featuring a dedicated signal conditioning and delta-sigma modulation integrated within a pixel area of 250 µm by 250 µm. The proof-of-concept receiver was implemented in an STMicroelectronics's 28-nm Fully Depleted Silicon On Insulator technology, and interfaces to a 4 × 4 subarray of capacitive micromachined ultrasound transducers (CMUTs). The front-end signal conditioning in each pixel employs a coarse/fine gain tuning architecture to fulfill the 90-dB dynamic range requirement of the application. The employed delta-sigma beamforming architecture obviates the need for area-consuming Nyquist ADCs and thereby enables an efficient in-pixel A/D conversion. The per-pixel switched-capacitor ΔΣ modulator leverages slewing-dominated and area-optimized inverter-based amplifiers. It occupies only 1/4th of the pixel, and its area compares favorably with state-of-the-art designs that offer the same SNR and bandwidth. The modulator's measured peak signal-to-noise-and-distortion ratio is 59.9 dB for a 10-MHz input bandwidth, and it consumes 6.65 mW from a 1-V supply. The overall subarray beamforming approach improves the area per channel by 7.4 times and the single-channel SNR by 8 dB compared to prior art with similar delay resolution and power dissipation. The functionality of the designed chip was evaluated within a PA imaging experiment, employing a flip-chip bonded 2-D CMUT array.

  View details for DOI 10.1109/JSSC.2017.2749425

  View details for PubMedID 31303662

  View details for PubMedCentralID PMC6625768

• A Pixel Pitch-Matched Ultrasound Receiver for 3-D Photoacoustic Imaging With Integrated Delta-Sigma Beamformer in 28-nm UTBB FD-SOI Chen, M., Perez, A., Kothapalli, S., Cathelin, P., Cathelin, A., Gambhir, S., Murmann, B. IEEE-INST ELECTRICAL ELECTRONICS ENGINEERS INC. 2017: 2843–56

  View details for DOI 10.1109/JSSC.2017.2749425

  View details for Web of Science ID 000413941800005

• SYNERGISTIC INHIBITION OF GLIOMA CELL PROLIFERATION BY WITHAFERIN A AND TUMOR TREATING FIELDS Chang, E., Pohling, C., Beygui, N., Patel, C. B., Rosenberg, J., Ha, D., Gambhir, S. S. OXFORD UNIV PRESS INC. 2017: 61–62

  View details for Web of Science ID 000415152501067

• Imaging of T cells in patients with recurrent glioblastoma TRANSLATIONAL CANCER RESEARCH Gambhir, S. 2017; 6: S291–S292

  View details for DOI 10.21037/tcr.2017.08.27

  View details for Web of Science ID 000413569400042

• F-FTC-146 in humans. Journal of nuclear medicine Hjørnevik, T., Cipriano, P. W., Shen, B., Hyung Park, J., Gulaka, P., Holley, D., Gandhi, H., Yoon, D., Mittra, E. S., Zaharchuk, G., Gambhir, S. S., McCurdy, C. R., Chin, F. T., Biswal, S. 2017

  Abstract

  The purpose of this study is to assess safety, biodistribution and radiation dosimetry in humans for the highly selective sigma-1 receptor (S1R) positron emission tomography (PET) agent (18)F-6-(3-fluoropropyl)-3-(2-(azepan-1-yl)ethyl)benzo[d]thiazol-2(3H)-one ((18)F-FTC-146). Methods: Ten healthy volunteers (HV; five female, five male; age: 34.3 ± 6.5 years) were recruited, and written informed consent was obtained from all participants. Series of whole-body PET/magnetic resonance imaging (PET/MRI) examinations were acquired for up to three hours after injection (357.2 ± 48.8 MBq). Blood samples were collected and standard vital signs (heart rate, pulse oximetry, and body temperature) were monitored at regular intervals. Regions-of-interest were delineated, time-activity curves were calculated, and organ uptake and dosimetry was estimated using PMOD 3.7 and Organ Linear Internal Dose Assessment (OLINDA). Results: All subjects tolerated the PET/MRI examination well, and no adverse reactions to (18)F-FTC-146 were reported. High accumulation of (18)F-FTC-146 was observed in S1R dense organs such as the pancreas and spleen, moderate uptake in the brain and myocardium, and low uptake in bone and muscle. High uptake was also observed in the kidneys and bladder, indicating renal tracer clearance. The effective dose (ED) of (18)F-FTC-146 was 0.0259 ± 0.0034 mSv/MBq (range: 0.0215-0.0301 mSv/MBq). Conclusion: First-in-human studies with clinical-grade (18)F-FTC-146 were successful. Injection of (18)F-FTC-146 is safe, and absorbed doses are acceptable. The potential of (18)F-FTC-146 as an imaging agent for a variety of neuroinflammatory diseases is currently under investigation.

  View details for DOI 10.2967/jnumed.117.192641

  View details for PubMedID 28572487

• A PET Imaging Strategy to Visualize Activated T Cells in Acute Graft-versus-Host Disease Elicited by Allogenic Hematopoietic Cell Transplant. Cancer research Ronald, J. A., Kim, B., Gowrishankar, G., Namavari, M., Alam, I. S., D'Souza, A., Nishikii, H., Chuang, H., Ilovich, O., Lin, C., Reeves, R., Shuhendler, A., Hoehne, A., Chan, C. T., Baker, J., Yaghoubi, S. S., VanBrocklin, H. F., Hawkins, R., Franc, B. L., Jivan, S., Slater, J. B., Verdin, E. F., Gao, K. T., Benjamin, J., Negrin, R., Gambhir, S. S. 2017; 77 (11): 2893-2902

  Abstract

  A major barrier to successful use of allogeneic hematopoietic cell transplantation is acute graft-versus-host disease (aGVHD), a devastating condition that arises when donor T cells attack host tissues. With current technologies, aGVHD diagnosis is typically made after end-organ injury and often requires invasive tests and tissue biopsies. This affects patient prognosis as treatments are dramatically less effective at late disease stages. Here, we show that a novel PET radiotracer, 2'-deoxy-2'-[18F]fluoro-9-β-D-arabinofuranosylguanine ([18F]F-AraG), targeted toward two salvage kinase pathways preferentially accumulates in activated primary T cells. [18F]F-AraG PET imaging of a murine aGVHD model enabled visualization of secondary lymphoid organs harboring activated donor T cells prior to clinical symptoms. Tracer biodistribution in healthy humans showed favorable kinetics. This new PET strategy has great potential for early aGVHD diagnosis, enabling timely treatments and improved patient outcomes. [18F]F-AraG may be useful for imaging activated T cells in various biomedical applications. Cancer Res; 77(11); 2893-902. ©2017 AACR.

  View details for DOI 10.1158/0008-5472.CAN-16-2953

  View details for PubMedID 28572504

• A Model-Based Personalized Cancer Screening Strategy for Detecting Early-Stage Tumors Using Blood-Borne Biomarkers CANCER RESEARCH Hori, S. S., Lutz, A. M., Paulmurugan, R., Gambhir, S. S. 2017; 77 (10): 2570-2584

  Abstract

  An effective cancer blood biomarker screening strategy must distinguish aggressive from non-aggressive tumors at an early, intervenable time. However, for blood-based strategies to be useful, the quality and quantiy of the biomarker shed into the blood and its relationship to tumor growth or progression must be validated. To study how blood biomarker levels correlate with early-stage viable tumor growth in an mouse model of human cancer, we monitored early tumor growth of engineered human ovarian cancer cells (A2780) implanted orthotopically into nude mice. Biomarker shedding was monitored by serial blood sampling, while tumor viability and volume was monitored by bioluminescence imaging and ultrasound imaging. From these metrics we developed a mathematical model of cancer biomarker kinetics in different compartments that accounts for biomarker shedding from tumor and healthy cells, biomarker entry into vasculature, biomarker elimination from plasma and subject-specific tumor growth. We validated the model in a separate set of mice where subject-specific tumor growth rates were accurately predicted. To illustrate clinical translation of this strategy, we allometrically scaled model parameters from mouse to human and used parameters for PSA shedding and prostate cancer. In this manner, we found that blood biomarker sampling data alone was capable of enabling the detection and discrimination of simulated aggressive (2-month tumor doubling time) and non-aggressive (18-month tumor doubling time) tumors as early as 7.2 months and 8.9 years before clinical imaging, respectively. Our model and screening strategy offer broad impact in their applicability to any solid cancer and the biomarkers they shed, thereby allowing a distinction between aggressive vs. non-aggressive tumors using blood biomarker sampling data alone.

  View details for DOI 10.1158/0008-5472.CAN-16-2904

  View details for Web of Science ID 000401252900003

  View details for PubMedID 28283654

• F-Fluoromaltotriose: A Second Generation PET Tracer Targeting the Maltodextrin Transporter in Bacteria. Journal of nuclear medicine Gowrishankar, G., Hardy, J., Wardak, M., Namavari, M., Reeves, R., Neofytou, E., Srinivasan, A., Wu, J., Contag, C., Gambhir, S. 2017

  Abstract

  Purpose: 6"-(18)F-fluoromaltotriose is a novel positron emission tomography (PET) tracer that can potentially be used to image and localize most bacterial infections, much like 2-deoxy-2-(18)F-fluoro-D-glucose ((18)F-FDG) has been used to image and localize many cancers. However, unlike (18)F-FDG, 6"-(18)F-fluoromaltotriose is not taken up by inflammatory lesions and appears to be specific to bacterial infections by targeting the maltodextrin transporter that is expressed in most Gram-positive and Gram-negative strains of bacteria. Materials and Methods: 6"-(18)F-fluoromaltotriose was synthesized with high radiochemical purity and evaluated in several clinically relevant bacterial strains incultures in vitro and in living mice. Results: 6"-(18)F-fluoromaltotriose was taken up in both Gram-positive and Gram-negative bacterial strains. 6"-[(18)F]-fluoromaltotriose was also able to detect Pseudomonas aeruginosa in a clinically relevant mouse model of wound infection. The utility of 6"-(18)F-fluoromaltotriose to help monitor antibiotic therapies was also evaluated in rats. Conclusion: 6"-(18)F-fluoromaltotriose is a promising new tracer that has significant diagnostic utility, with the potential to change the clinical management of patients suffering from infectious diseases of bacterial origin.

  View details for DOI 10.2967/jnumed.117.191452

  View details for PubMedID 28490473

• Initial Experience with a New PET/CT System Using SiPM Detectors: Image Quality Comparison with Standard PET/CT Park, S., Baratto, L., Hatami, N., Davidzon, G., Srinivas, S., Gambhir, S., Lagaru, A. SOC NUCLEAR MEDICINE INC. 2017

  View details for Web of Science ID 000404949906122

• Towards clinically translatable in vivo nanodiagnostics. Nature reviews. Materials Park, S. M., Aalipour, A., Vermesh, O., Yu, J. H., Gambhir, S. S. 2017; 2 (5)

  Abstract

  Nanodiagnostics as a field makes use of fundamental advances in nanobiotechnology to diagnose, characterize and manage disease at the molecular scale. As these strategies move closer to routine clinical use, a proper understanding of different imaging modalities, relevant biological systems and physical properties governing nanoscale interactions is necessary to rationally engineer next-generation bionanomaterials. In this Review, we analyse the background physics of several clinically relevant imaging modalities and their associated sensitivity and specificity, provide an overview of the materials currently used for in vivo nanodiagnostics, and assess the progress made towards clinical translation. This work provides a framework for understanding both the impressive progress made thus far in the nanodiagnostics field as well as presenting challenges that must be overcome to obtain widespread clinical adoption.

  View details for DOI 10.1038/natrevmats.2017.14

  View details for PubMedID 29876137

  View details for PubMedCentralID PMC5985817

• Regulatory Aspects of Optical Methods and Exogenous Targets for Cancer Detection CANCER RESEARCH Tummers, W. S., Warram, J. M., Tipirneni, K. E., Fengler, J., Jacobs, P., Shankar, L., Henderson, L., Ballard, B., Pogue, B. W., Weichert, J. P., Bouvet, M., Sorger, J., Contag, C. H., Frangioni, J. V., Tweedle, M. F., Basilion, J. P., Gambhir, S. S., Rosenthal, E. L. 2017; 77 (9): 2197-2206

  Abstract

  Considerable advances in cancer-specific optical imaging have improved the precision of tumor resection. In comparison to traditional imaging modalities, this technology is unique in its ability to provide real-time feedback to the operating surgeon. Given the significant clinical implications of optical imaging, there is an urgent need to standardize surgical navigation tools and contrast agents to facilitate swift regulatory approval. Because fluorescence-enhanced surgery requires a combination of both device and drug, each may be developed in conjunction, or separately, which are important considerations in the approval process. This report is the result of a one-day meeting held on May 4, 2016 with officials from the National Cancer Institute, the FDA, members of the American Society of Image-Guided Surgery, and members of the World Molecular Imaging Society, which discussed consensus methods for FDA-directed human testing and approval of investigational optical imaging devices as well as contrast agents for surgical applications. The goal of this workshop was to discuss FDA approval requirements and the expectations for approval of these novel drugs and devices, packaged separately or in combination, within the context of optical surgical navigation. In addition, the workshop acted to provide clarity to the research community on data collection and trial design. Reported here are the specific discussion items and recommendations from this critical and timely meeting. Cancer Res; 77(9); 2197-206. ©2017 AACR.

  View details for DOI 10.1158/0008-5472.CAN-16-3217

  View details for Web of Science ID 000400270100004

  View details for PubMedID 28428283

• Harnessing Radioluminescence and Sound to Reveal Molecular Pathology of Atherosclerotic Plaques Zaman, R., Yousefi, S., Long, S., Contag, C., Gambhir, S., Khuri-Yakub, B., Xing, L. SOC NUCLEAR MEDICINE INC. 2017

  View details for Web of Science ID 000404949900032

• SiPM PET/CT vs. Standard PET/CT: A Pilot Study Comparing Semi-Quantitative Measurements in Normal Tissues and Lesions Baratto, L., Park, S., Hatami, N., Davidzon, G., Srinivas, S., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2017

  View details for Web of Science ID 000404949902041

• Natural product-inspired agents and their anticancer activity against glioblastoma multiforme cells Kumar, V., Banister, S., Kaufman, H., Vittimberga, J., Jacobo, S., Gambhir, S., Malhotra, S. AMER CHEMICAL SOC. 2017

  View details for Web of Science ID 000430569104415

• Practical Immuno-PET Radiotracer Design Considerations for Human Immune Checkpoint Imaging JOURNAL OF NUCLEAR MEDICINE Mayer, A. T., Natarajan, A., Gordon, S. R., Maute, R. L., McCracken, M. N., Ring, A. M., Weissman, I. L., Gambhir, S. S. 2017; 58 (4): 538-546

  Abstract

  Immune checkpoint blockade has emerged as a promising cancer treatment paradigm. Unfortunately, there are still a large number of patients and malignancies that do not respond to therapy. A major barrier to validating biomarkers for the prediction and monitoring of responders to clinical checkpoint blockade has been the lack of imaging tools to accurately assess dynamic immune checkpoint expression. Here, we sought to optimize noninvasive immuno-PET imaging of human programmed death-ligand 1 (PD-L1) expression, in a preclinical model, using a small high-affinity engineered protein scaffold (HAC-PD1). Six HAC-PD1 radiotracer variants were developed and used in preclinical imaging and biodistribution studies to assess their ability to detect human PD-L1 expression in vivo. Radiotracer design modifications included chelate, glycosylation, and radiometal. HACA-PD1 was adopted as the naming convention for aglycosylated tracer variants. NOD scid γ-(NSG) mice were inoculated with subcutaneous tumors engineered to either be constitutively positive (CT26 hPD-L1) or be negative (ΔmPD-L1 CT26) for human PD-L1 expression. When the tumors had grown to an average size of 1 cm in diameter, mice were injected with 0.75-2.25 MBq (∼10 μg) of an engineered radiotracer variant and imaged. At 1 h after injection, organs were harvested for biodistribution. Of the practical immuno-PET tracer modifications considered, glycosylation was the most prominent design factor affecting tracer uptake, specificity, and clearance. In imaging studies, aglycosylated (64)Cu-NOTA-HACA-PD1 most accurately visualized human PD-L1 expression in vivo. We reasoned that because of the scaffold's small size (14 kDa), its pharmacokinetics may be suitable for labeling with the short-lived and widely clinically available radiometal (68)Ga. At 1 h after injection, (68)Ga-NOTA-HACA-PD1 and (68)Ga-DOTA-HACA-PD1 exhibited promising target-to-background ratios in ex vivo biodistribution studies (12.3 and 15.2 tumor-to-muscle ratios, respectively). Notably, all HAC-PD1 radiotracer variants enabled much earlier detection of human PD-L1 expression (1 h after injection) than previously reported radiolabeled antibodies (>24 h after injection). This work provides a template for assessing immuno-PET tracer design parameters and supports the translation of small engineered protein radiotracers for imaging human immune checkpoints.

  View details for DOI 10.2967/jnumed.116.177659

  View details for Web of Science ID 000398249600012

  View details for PubMedCentralID PMC5373501

• Ultrasound Molecular Imaging With BR55 in Patients With Breast and Ovarian Lesions: First-in-Human Results. Journal of clinical oncology Willmann, J. K., Bonomo, L., Carla Testa, A., Rinaldi, P., Rindi, G., Valluru, K. S., Petrone, G., Martini, M., Lutz, A. M., Gambhir, S. S. 2017: JCO2016708594-?

  Abstract

  Purpose We performed a first-in-human clinical trial on ultrasound molecular imaging (USMI) in patients with breast and ovarian lesions using a clinical-grade contrast agent (kinase insert domain receptor [KDR] -targeted contrast microbubble [MBKDR]) that is targeted at the KDR, one of the key regulators of neoangiogenesis in cancer. The aim of this study was to assess whether USMI using MBKDR is safe and allows assessment of KDR expression using immunohistochemistry (IHC) as the gold standard. Methods Twenty-four women (age 48 to 79 years) with focal ovarian lesions and 21 women (age 34 to 66 years) with focal breast lesions were injected intravenously with MBKDR (0.03 to 0.08 mL/kg of body weight), and USMI of the lesions was performed starting 5 minutes after injection up to 29 minutes. Blood pressure, ECG, oxygen levels, heart rate, CBC, and metabolic panel were obtained before and after MBKDR administration. Persistent focal MBKDR binding on USMI was assessed. Patients underwent surgical resection of the target lesions, and tissues were stained for CD31 and KDR by IHC. Results USMI with MBKDR was well tolerated by all patients without safety concerns. Among the 40 patients included in the analysis, KDR expression on IHC matched well with imaging signal on USMI in 93% of breast and 85% of ovarian malignant lesions. Strong KDR-targeted USMI signal was present in 77% of malignant ovarian lesions, with no targeted signal seen in 78% of benign ovarian lesions. Similarly, strong targeted signal was seen in 93% of malignant breast lesions with no targeted signal present in 67% of benign breast lesions. Conclusion USMI with MBKDR is clinically feasible and safe, and KDR-targeted USMI signal matches well with KDR expression on IHC. This study lays the foundation for a new field of clinical USMI in cancer.

  View details for DOI 10.1200/JCO.2016.70.8594

  View details for PubMedID 28291391

• F]FTC-146. Molecular imaging and biology Shen, B., Park, J. H., Hjørnevik, T., Cipriano, P. W., Yoon, D., Gulaka, P. K., Holly, D., Behera, D., Avery, B. A., Gambhir, S. S., McCurdy, C. R., Biswal, S., Chin, F. T. 2017

  Abstract

  Sigma-1 receptors (S1Rs) play an important role in many neurological disorders. Simultaneous positron emission tomography (PET)/magnetic resonance imaging (MRI) with S1R radioligands may provide valuable information for diagnosing and guiding treatment for these diseases. Our previously reported S1R radioligand, [(18)F]FTC-146, demonstrated high affinity for the S1R (K i = 0.0025 nM) and excellent selectivity for the S1R over the sigma-2 receptor (S2Rs; K i = 364 nM) across several species (from mouse to non-human primate). Herein, we report the clinical-grade radiochemistry filed with exploratory Investigational New Drug (eIND) and first-in-human PET/MRI evaluation of [(18)F]FTC-146.[(18)F]FTC-146 is prepared via a direct [(18)F] fluoride nucleophilic radiolabeling reaction and formulated in 0.9 % NaCl containing no more than 10 % ethanol through sterile filtration. Quality control (QC) was performed based on USP 823 before doses were released for clinical use. The safety and whole body biodistribution of [(18)F]FTC-146 were evaluated using a simultaneous PET/MR scanner in two representative healthy human subjects.[(18)F]FTC-146 was synthesized with a radiochemical yield of 3.3 ± 0.7 % and specific radioactivity of 8.3 ± 3.3 Ci/μmol (n = 10, decay corrected to EOB). Both radiochemical and chemical purities were >95 %; the prepared doses were stable for 4 h at ambient temperature. All QC test results met specified clinical criteria. The in vivo PET/MRI investigations showed that [(18)F]FTC-146 rapidly crossed the blood brain barrier and accumulated in S1R-rich regions of the brain. There were also radioactivity distributed in the peripheral organs, i.e., the lungs, spleen, pancreas, and thyroid. Furthermore, insignificant uptake of [(18)F]FTC-146 was observed in cortical bone and muscle.A reliable and automated radiosynthesis for providing routine clinical-grade [(18)F]FTC-146 for human studies was established in a modified GE TRACERlab FXFN. PET/MRI demonstrated the initial tracer biodistribution in humans, and clinical studies investigating different S1R-related diseases are in progress.

  View details for DOI 10.1007/s11307-017-1064-z

  View details for PubMedID 28280965

• Development of Novel ImmunoPET Tracers to Image Human PD-1 Checkpoint Expression on Tumor-Infiltrating Lymphocytes in a Humanized Mouse Model. Molecular imaging and biology Natarajan, A., Mayer, A. T., Reeves, R. E., Nagamine, C. M., Gambhir, S. S. 2017

  Abstract

  It is well known that cancers exploit immune checkpoints (programmed death 1 receptor (PD-1) and its ligand (PD-L1)) to evade anti-tumor immune responses. Although immune checkpoint (IC) blockade is a promising approach, not all patients respond. Hence, imaging of tumor-infiltrating lymphocytes (TILs) is of high specific interest, as they are known to express PD-1 during activation and subsequent exhaustion in the tumor microenvironment and are thought to be potentially predictive of therapeutic responses to IC blockade.We developed immune-tracers for positron emission tomography (PET) to image hPD-1 status of human peripheral blood mononuclear cells (hPBMCs) adoptively transferred to NOD-scid IL-2Rγ(null) (NSG) mice (hNSG) bearing A375 human skin melanoma tumors. The anti-PD-1 human antibody (IgG; keytruda) was labeled with either Zr-89 or Cu-64 radiometals to image PD-1-expressing human TILs in vivo.[(89)Zr] Keytruda (groups = 2; NSG-ctl (control) and hNSG-nblk (non-blocking), n = 3-5, 3.2 ± 0.4 MBq/15-16 μg/200 μl) and [(64)Cu] Keytruda (groups = 3; NSG-ctl, NSG-blk (blocking), and hNSG-nblk; n = 4, 7.4 ± 0.4 MBq /20-25 μg/200 μl) were administered in mice. PET-CT scans were performed over 1-144 h ([(89)Zr] Keytruda) and 1-48 h ([(64)Cu] Keytruda) on mice. hNSG mice exhibited a high tracer uptake in the spleen, lymphoid organs and tumors. At 24 h, human TILs homing into melanoma of hNSG-nblk mice exhibited high signal (mean %ID/g ± SD) of 3.8 ± 0.4 ([(89)Zr] Keytruda), and 6.4 ± 0.7 ([(64)Cu] Keytruda), which was 1.5- and 3-fold higher uptake compared to NSG-ctl mice (p = 0.01), respectively. Biodistribution measurements of hNSG-nblk mice performed at 144 h ([(89)Zr] Keytruda) and 48 h ([(64)Cu] Keytruda) p.i. revealed tumor to muscle ratios as high as 45- and 12-fold, respectively.Our immunoPET study clearly demonstrates specific imaging of human PD-1-expressing TILs within the tumor and lymphoid tissues. This suggests these anti-human-PD-1 tracers could be clinically translatable to monitor cancer treatment response to IC blockade therapy.

  View details for DOI 10.1007/s11307-017-1060-3

  View details for PubMedID 28247187

• F]DASA-23 for Imaging Tumor Glycolysis Through Noninvasive Measurement of Pyruvate Kinase M2. Molecular imaging and biology Beinat, C., Alam, I. S., James, M. L., Srinivasan, A., Gambhir, S. S. 2017

  Abstract

  A hallmark of cancer is metabolic reprogramming, which is exploited by cancer cells to ensure rapid growth and survival. Pyruvate kinase M2 (PKM2) catalyzes the final step in glycolysis, a key step in tumor metabolism and growth. Recently, we reported the radiosynthesis of the first positron emission tomography tracer for visualizing PKM2 in vivo-i.e., [(11)C]DASA-23. Due to the highly promising imaging results obtained with [(11)C]DASA-23 in rodent model glioblastoma, we set out to generate an F-18-labeled version of this tracer, with the end goal of clinical translation in mind. Herein, we report the radiosynthesis of 1-((2-fluoro-6-[(18)F]fluorophenyl)sulfonyl)-4-((4-methoxyphenyl)sulfonyl)piperazine ([(18)F]DASA-23) and our initial investigation of its binding properties in cancer cells.We synthesized [(18)F]DASA-23 via fluorination of 1-((2-fluoro-6-nitrophenyl)sulfonyl)-4-((4-methoxyphenyl)sulfonyl)piperazine (10) with K[(18)F]F/K2.2.2 in N,N-dimethylformamide at 110 °C for 20 min. Subsequently, we evaluated uptake of [(18)F]DASA-23 in HeLa cervical adenocarcinoma cells and in vitro stability in human and mouse serum.We successfully prepared [(18)F]DASA-23 in 2.61 ± 1.54 % radiochemical yield (n = 10, non-decay corrected at end of synthesis) with a specific activity of 2.59 ± 0.44 Ci/μmol. Preliminary cell uptake experiments revealed high uptake in HeLa cells, which was effectively blocked by pretreating cells with the structurally distinct PKM2 activator, TEPP-46. [(18)F]DASA-23 remained intact in human and mouse serum up to 120 min.Herein, we have identified a F-18-labeled PKM2 specific radiotracer which shows potential for in vivo imaging. The promising cell uptake results reported herein warrant the further evaluation of [(18)F]DASA-23 for its ability to detect and monitor cancer noninvasively.

  View details for DOI 10.1007/s11307-017-1068-8

  View details for PubMedID 28236227

• Nanomaterials for In Vivo Imaging. Chemical reviews Smith, B. R., Gambhir, S. S. 2017; 117 (3): 901-986

  Abstract

  In vivo imaging, which enables us to peer deeply within living subjects, is producing tremendous opportunities both for clinical diagnostics and as a research tool. Contrast material is often required to clearly visualize the functional architecture of physiological structures. Recent advances in nanomaterials are becoming pivotal to generate the high-resolution, high-contrast images needed for accurate, precision diagnostics. Nanomaterials are playing major roles in imaging by delivering large imaging payloads, yielding improved sensitivity, multiplexing capacity, and modularity of design. Indeed, for several imaging modalities, nanomaterials are now not simply ancillary contrast entities, but are instead the original and sole source of image signal that make possible the modality's existence. We address the physicochemical makeup/design of nanomaterials through the lens of the physical properties that produce contrast signal for the cognate imaging modality-we stratify nanomaterials on the basis of their (i) magnetic, (ii) optical, (iii) acoustic, and/or (iv) nuclear properties. We evaluate them for their ability to provide relevant information under preclinical and clinical circumstances, their in vivo safety profiles (which are being incorporated into their chemical design), their modularity in being fused to create multimodal nanomaterials (spanning multiple different physical imaging modalities and therapeutic/theranostic capabilities), their key properties, and critically their likelihood to be clinically translated.

  View details for DOI 10.1021/acs.chemrev.6b00073

  View details for PubMedID 28045253

• Detection of Stem Cell Transplant Rejection with Ferumoxytol MR Imaging: Correlation of MR Imaging Findings with Those at Intravital Microscopy. Radiology Daldrup-Link, H. E., Chan, C., Lenkov, O., Taghavigarmestani, S., Nazekati, T., Nejadnik, H., Chapelin, F., Khurana, A., Tong, X., Yang, F., Pisani, L., Longaker, M., Gambhir, S. S. 2017: 161139-?

  Abstract

  Purpose To determine whether endogenous labeling of macrophages with clinically applicable nanoparticles enables noninvasive detection of innate immune responses to stem cell transplants with magnetic resonance (MR) imaging. Materials and Methods Work with human stem cells was approved by the institutional review board and the stem cell research oversight committee, and animal experiments were approved by the administrative panel on laboratory animal care. Nine immunocompetent Sprague-Dawley rats received intravenous injection of ferumoxytol, and 18 Jax C57BL/6-Tg (Csf1r-EGFP-NGFR/FKBP1A/TNFRSF6) 2Bck/J mice received rhodamine-conjugated ferumoxytol. Then, 48 hours later, immune-matched or mismatched stem cells were implanted into osteochondral defects of the knee joints of experimental rats and calvarial defects of Jax mice. All animals underwent serial MR imaging and intravital microscopy (IVM) up to 4 weeks after surgery. Macrophages of Jax C57BL/6-Tg (Csf1r-EGFP-NGFR/FKBP1A/TNFRSF6) 2Bck/J mice express enhanced green fluorescent protein (GFP), which enables in vivo correlation of ferumoxytol enhancement at MR imaging with macrophage quantities at IVM. All quantitative data were compared between experimental groups by using a mixed linear model and t tests. Results Immune-mismatched stem cell implants demonstrated stronger ferumoxytol enhancement than did matched stem cell implants. At 4 weeks, T2 values of mismatched implants were significantly lower than those of matched implants in osteochondral defects of female rats (mean, 10.72 msec for human stem cells and 11.55 msec for male rat stem cells vs 15.45 msec for sex-matched rat stem cells; P = .02 and P = .04, respectively) and calvarial defects of recipient mice (mean, 21.7 msec vs 27.1 msec, respectively; P = .0444). This corresponded to increased recruitment of enhanced GFP- and rhodamine-ferumoxytol-positive macrophages into stem cell transplants, as visualized with IVM and histopathologic examination. Conclusion Endogenous labeling of macrophages with ferumoxytol enables noninvasive detection of innate immune responses to stem cell transplants with MR imaging. (©) RSNA, 2017 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.2017161139

  View details for PubMedID 28128708

• Reporter gene imaging of targeted T cell immunotherapy in recurrent glioma. Science translational medicine Keu, K. V., Witney, T. H., Yaghoubi, S., Rosenberg, J., Kurien, A., Magnusson, R., Williams, J., Habte, F., Wagner, J. R., Forman, S., Brown, C., Allen-Auerbach, M., Czernin, J., Tang, W., Jensen, M. C., Badie, B., Gambhir, S. S. 2017; 9 (373)

  Abstract

  High-grade gliomas are aggressive cancers that often become rapidly fatal. Immunotherapy using CD8(+) cytotoxic T lymphocytes (CTLs), engineered to express both herpes simplex virus type 1 thymidine kinase (HSV1-TK) and interleukin-13 (IL-13) zetakine chimeric antigen receptor (CAR), is a treatment strategy with considerable potential. To optimize this and related immunotherapies, it would be helpful to monitor CTL viability and trafficking to glioma cells. We show that noninvasive positron emission tomography (PET) imaging with 9-[4-[(18)F]fluoro-3-(hydroxymethyl)butyl]guanine ([(18)F]FHBG) can track HSV1-tk reporter gene expression present in CAR-engineered CTLs. [(18)F]FHBG imaging was safe and enabled the longitudinal imaging of T cells stably transfected with a PET reporter gene in patients. Further optimization of this imaging approach for monitoring in vivo cell trafficking should greatly benefit various cell-based therapies for cancer.

  View details for DOI 10.1126/scitranslmed.aag2196

  View details for PubMedID 28100832

  View details for PubMedCentralID PMC5260938

• 18F-FDG silicon photomultiplier PET/CT: A pilot study comparing semi-quantitative measurements with standard PET/CT. PloS one Baratto, L., Park, S. Y., Hatami, N., Davidzon, G., Srinivas, S., Gambhir, S. S., Iagaru, A. 2017; 12 (6)

  Abstract

  To evaluate if the new Discovery Molecular Insights (DMI) PET/CT scanner provides equivalent results compared to the standard of care PET/CT scanners (GE Discovery 600 or GE Discovery 690) used in our clinic and to explore any possible differences in semi-quantitative measurements.The local Institutional Review Board approved the protocol and written informed consent was obtained from each patient. Between September and November 2016, 50 patients underwent a single 18F-FDG injection and two scans: the clinical standard PET/CT followed immediately by the DMI PET/CT scan. We measured SUVmax and SUVmean of different background organs and up to four lesions per patient from data acquired using both scanners.DMI PET/CT identified all the 107 lesions detected by standard PET/CT scanners, as well as additional 37 areas of focal increased 18F-FDG uptake. The SUVmax values for all 107 lesions ranged 1.2 to 14.6 (mean ± SD: 2.8 ± 2.8), higher on DMI PET/CT compared with standard of care PET/CT. The mean lesion:aortic arch SUVmax ratio and mean lesion:liver SUVmax ratio were 0.2-15.2 (mean ± SD: 3.2 ± 2.6) and 0.2-8.5 (mean ± SD: 1.9 ± 1.4) respectively, higher on DMI PET/CT than standard PET/CT. These differences were statistically significant (P value < 0.0001) and not correlated to the delay in acquisition of DMI PET data (P < 0.0001).Our study shows high performance of the new DMI PET/CT scanner. This may have a significant role in diagnosing and staging disease, as well as for assessing and monitoring responses to therapies.

  View details for DOI 10.1371/journal.pone.0178936

  View details for PubMedID 28582472

• Imaging B-Cells in a Mouse of Multiple Sclerosis Using 64Cu-Rituximab-PET Journal of Nuclear Medicine James, M. L., Hoehne, A., Mayer, A. T., Lechtenburg, K., Moreno, M., Gowrishankar, G., Illovich, O., Natarajan, A., Johnson, E. M., Nguyen , J., Quach, L., Han, M., Buckwalter, M., Chandra, S., Gambhir, S. S. 2017; 58 (11): 1845-1851
• Capture and Genetic Analysis of Circulating Tumor Cells Using a Magnetic Separation Device (Magnetic Sifter) CIRCULATING TUMOR CELLS: METHODS AND PROTOCOLS Ooi, C., Park, S., Wong, D. J., Gambhir, S. S., Wang, S. X., Magbanua, M. J., Park, J. W. 2017; 1634: 153–62

  View details for DOI 10.1007/978-1-4939-7144-2_12

  View details for Web of Science ID 000449398600014

• On-Target Probes for Early Detection Nature Biomedical Engineering Hori, S. S., Tummers, W. S., Gambhir, S. S. 2017; 1: 1-3

  View details for DOI 10.1038/s41551-017-0062

• Towards clinically translatable in vivo nanodiagnostics Nature Reviews Materials Park, S., Aalipour, A., Vermesh, O., Yu, J., Gambhir, S. S. 2017; 2

  View details for DOI 10.1038/natrevmats.2017.14

• Multigene Profiling of Single Circulating Tumor Cells Molecular & Cellular Oncology Park, S., Wong, D., Ooi, C., Nesvet, J., Nair, V. S., Wang, S. X., Gambhir, S. S. 2017; 4 (2): e1289295

  Abstract

  Numerous techniques for isolating circulating tumor cells (CTCs) have been developed. Concurrently, single-cell techniques that can reveal molecular components of CTCs have become widely available. We discuss how the combination of isolation and multigene profiling of single CTCs in our platform can facilitate eventual translation to the clinic.

  View details for DOI 10.1080/23723556.2017.1289295

  View details for PubMedCentralID PMC5383366

• High-throughput full-length single-cell mRNA-seq of rare cells. PloS one Ooi, C. C., Mantalas, G. L., Koh, W. n., Neff, N. F., Fuchigami, T. n., Wong, D. J., Wilson, R. J., Park, S. M., Gambhir, S. S., Quake, S. R., Wang, S. X. 2017; 12 (11): e0188510

  Abstract

  Single-cell characterization techniques, such as mRNA-seq, have been applied to a diverse range of applications in cancer biology, yielding great insight into mechanisms leading to therapy resistance and tumor clonality. While single-cell techniques can yield a wealth of information, a common bottleneck is the lack of throughput, with many current processing methods being limited to the analysis of small volumes of single cell suspensions with cell densities on the order of 107 per mL. In this work, we present a high-throughput full-length mRNA-seq protocol incorporating a magnetic sifter and magnetic nanoparticle-antibody conjugates for rare cell enrichment, and Smart-seq2 chemistry for sequencing. We evaluate the efficiency and quality of this protocol with a simulated circulating tumor cell system, whereby non-small-cell lung cancer cell lines (NCI-H1650 and NCI-H1975) are spiked into whole blood, before being enriched for single-cell mRNA-seq by EpCAM-functionalized magnetic nanoparticles and the magnetic sifter. We obtain high efficiency (> 90%) capture and release of these simulated rare cells via the magnetic sifter, with reproducible transcriptome data. In addition, while mRNA-seq data is typically only used for gene expression analysis of transcriptomic data, we demonstrate the use of full-length mRNA-seq chemistries like Smart-seq2 to facilitate variant analysis of expressed genes. This enables the use of mRNA-seq data for differentiating cells in a heterogeneous population by both their phenotypic and variant profile. In a simulated heterogeneous mixture of circulating tumor cells in whole blood, we utilize this high-throughput protocol to differentiate these heterogeneous cells by both their phenotype (lung cancer versus white blood cells), and mutational profile (H1650 versus H1975 cells), in a single sequencing run. This high-throughput method can help facilitate single-cell analysis of rare cell populations, such as circulating tumor or endothelial cells, with demonstrably high-quality transcriptomic data.

  View details for PubMedID 29186152

  View details for PubMedCentralID PMC5706670

• Fdg Pet-CT Suvmax And Circulating Tumor Microemboli Identify Recurrence In Patients With Non-Small Cell Lung Cancer Nair, V. S., Carlsson, F., Carlsson, A., Jamali, M., Keu, K., Vasanawala, M., Shrager, J., Loo, B. W., Horng, G., Kuschner, W., Gambhir, S. S., Kuhn, P. AMER THORACIC SOC. 2017

  View details for Web of Science ID 000400372500266

• Capture and Genetic Analysis of Circulating Tumor Cells Using a Magnetic Separation Device (Magnetic Sifter). Methods in molecular biology (Clifton, N.J.) Ooi, C. C., Park, S. M., Wong, D. J., Gambhir, S. S., Wang, S. X. 2017; 1634: 153–62

  Abstract

  Circulating tumor cells (CTCs) are currently widely studied for their potential application as part of a liquid biopsy. These cells are shed from the primary tumor into the circulation, and are postulated to provide insight into the molecular makeup of the actual tumor in a minimally invasive manner. However, they are extremely rare in blood, with typical concentrations of 1-100 in a milliliter of blood; hence, a need exists for a rapid and high-purity method for isolating CTCs from whole blood. Here, we describe the application of a microfabricated magnetic sifter toward isolation of CTCs from whole blood at volumetric flow rates of 10 mL/h, along with the use of a PDMS-based nanowell system for single-cell gene expression profiling. This method allows rapid isolation of CTCs and subsequent integration with downstream genetic profiling methods for clinical applications such as targeted therapy, therapy monitoring, or further biological studies.

  View details for PubMedID 28819848

• Synergistic inhibition of glioma cell proliferation by Withaferin A and tumor treating fields. Journal of neuro-oncology Chang, E. n., Pohling, C. n., Beygui, N. n., Patel, C. B., Rosenberg, J. n., Ha, D. H., Gambhir, S. S. 2017

  Abstract

  Glioblastoma (GBM) is the most aggressive and lethal form of brain cancer. Standard therapies are non-specific and often of limited effectiveness; thus, efforts are underway to uncover novel, unorthodox therapies against GBM. In previous studies, we investigated Withaferin A, a steroidal lactone from Ayurvedic medicine that inhibits proliferation in cancers including GBM. Another novel approach, tumor treating fields (TTFields), is thought to disrupt mitotic spindle formation and stymie proliferation of actively dividing cells. We hypothesized that combining TTFields with Withaferin A would synergistically inhibit proliferation in glioblastoma. Human glioblastoma cells (GBM2, GBM39, U87-MG) and human breast adenocarcinoma cells (MDA-MB-231) were isolated from primary tumors. The glioma cell lines were genetically engineered to express firefly luciferase. Proliferative potential was assessed either by bioluminescence imaging or cell counting via hemocytometer. TTFields (4 V/cm) significantly inhibited growth of the four cancer cell lines tested (n = 3 experiments per time point, four measurements per sample, p < 0.02 at least; 2-way ANOVA, control vs. treatment). The combination of Withaferin A (10-100 nM) with TTFields significantly inhibited the growth of the glioma cells to a degree beyond that of Withaferin A or TTFields alone. The interaction of the Withaferin A and TTFields on glioma cells was found to be synergistic in nature (p < 0.01, n = 3 experiments). These findings were validated by both bioluminescence and hemocytometric measurements. The combination of Withaferin A with TTFields represents a novel approach to treat GBM in a manner that is likely better than either treatment alone and that is synergistic.

  View details for PubMedID 28681243

• A novel theranostic strategy for MMP-14 expressing glioblastomas impacts survival. Molecular cancer therapeutics Mohanty, S. n., Chen, Z. n., Li, K. n., Morais, G. R., Klockow, J. n., Yerneni, K. n., Pisani, L. n., Chin, F. T., Mitra, S. n., Cheshier, S. n., Chang, E. n., Gambhir, S. S., Rao, J. n., Loadman, P. M., Falconer, R. A., Daldrup-Link, H. E. 2017

  Abstract

  Glioblastoma (GBM) has a dismal prognosis. Evidence from preclinical tumor models and human trials indicates the role of GBM initiating cells (GIC) in GBM drug resistance. Here, we propose a new treatment option with tumor enzyme-activatable, combined therapeutic and diagnostic (theranostic) nanoparticles, which caused specific toxicity against GBM tumor cells and GICs. The theranostic cross-linked iron oxide nanoparticles (CLIO) were conjugated to a highly potent vascular disrupting agent (ICT) and secured with a matrix-metalloproteinase (MMP-14) cleavable peptide. Treatment with CLIO-ICT disrupted tumor vasculature of MMP-14 expressing GBM, induced GIC apoptosis and significantly impaired tumor growth. In addition, the iron core of CLIO-ICT enabled in vivo drug tracking with MR imaging. Treatment with CLIO-ICT plus temozolomide achieved tumor remission and significantly increased survival of human GBM bearing mice by more than 2 fold compared to treatment with temozolomide alone. Thus, we present a novel therapeutic strategy with significant impact on survival and great potential for clinical translation.

  View details for PubMedID 28659432

• Imaging B cells in a mouse model of multiple sclerosis using (64)Cu-Rituximab-PET. Journal of nuclear medicine : official publication, Society of Nuclear Medicine James, M. L., Hoehne, A. n., Mayer, A. T., Lechtenberg, K. n., Moreno, M. n., Gowrishankar, G. n., Ilovich, O. n., Natarajan, A. n., Johnson, E. M., Nguyen, J. n., Quach, L. n., Han, M. n., Buckwalter, M. n., Chandra, S. n., Gambhir, S. S. 2017

  Abstract

  B lymphocytes are a key pathological feature of multiple sclerosis (MS), and are becoming an important therapeutic target for this condition. Currently, there is no approved technique to non-invasively visualize B cells in the central nervous system (CNS) to monitor MS disease progression and response to therapies. Here we evaluated (64)Cu-Rituximab, a radiolabeled antibody specifically targeting the human B cell marker CD20, for its ability to image B cells in a mouse model of MS using positron emission tomography (PET). Methods: To model CNS infiltration by B cells, experimental autoimmune encephalomyelitis (EAE) was induced in transgenic mice that express human CD20 on B cells. EAE mice were given subcutaneous injections of Myelin Oligodendrocyte Glycoprotein fragment1-125 (MOG1-125) emulsified in complete Freund's adjuvant. Control mice received complete Freund's adjuvant alone. PET imaging of EAE and control mice was performed 1, 4, and 19h following (64)Cu-Rituximab administration. Mice were perfused and sacrificed after final PET scan, and radioactivity in dissected tissues was measured with a gamma-counter. CNS tissues from these mice were immunostained to quantify B cells or further analyzed via digital autoradiography. Results: Lumbar spinal cord PET signal was significantly higher in EAE mice compared to controls at all evaluated time points (e.g., 1h post-injection: 5.44 ± 0.37 vs. 3.33 ± 0.20 %ID/g, p<0.05). (64)Cu-Rituximab-PET signal in brain regions ranged between 1.74 ± 0.11 and 2.93 ± 0.15 %ID/g for EAE mice compared to 1.25±0.08 and 2.24±0.11%ID/g for controls, p<0.05 for all regions except striatum and thalamus at 1h post-injection. Similarly, ex vivo biodistribution results revealed notably higher (64)Cu-Rituximab uptake in brain and spinal cord of huCD20tg EAE, and B220 immunostaining verified that increased (64)Cu-Rituximab uptake in CNS tissues corresponded with elevated B cells. Conclusion: B cells can be detected in the CNS of EAE mice using (64)Cu-Rituximab-PET. Results from these studies warrant further investigation of (64)Cu-Rituximab in EAE models and consideration of use in MS patients to evaluate its potential for detecting and monitoring B cells in the progression and treatment of this disease. These results represent an initial step toward generating a platform to evaluate B cell-targeted therapeutics en route to the clinic.

  View details for PubMedID 28687602

• The Exosome Total Isolation Chip. ACS nano Liu, F. n., Vermesh, O. n., Mani, V. n., Ge, T. J., Madsen, S. J., Sabour, A. n., Hsu, E. C., Gowrishankar, G. n., Kanada, M. n., Jokerst, J. V., Sierra, R. G., Chang, E. n., Lau, K. n., Sridhar, K. n., Bermudez, A. n., Pitteri, S. J., Stoyanova, T. n., Sinclair, R. n., Nair, V. S., Gambhir, S. S., Demirci, U. n. 2017

  Abstract

  Circulating tumor-derived extracellular vesicles (EVs) have emerged as a promising source for identifying cancer biomarkers for early cancer detection. However, the clinical utility of EVs has thus far been limited by the fact that most EV isolation methods are tedious, nonstandardized, and require bulky instrumentation such as ultracentrifugation (UC). Here, we report a size-based EV isolation tool called ExoTIC (exosome total isolation chip), which is simple, easy-to-use, modular, and facilitates high-yield and high-purity EV isolation from biofluids. ExoTIC achieves an EV yield ∼4-1000-fold higher than that with UC, and EV-derived protein and microRNA levels are well-correlated between the two methods. Moreover, we demonstrate that ExoTIC is a modular platform that can sort a heterogeneous population of cancer cell line EVs based on size. Further, we utilize ExoTIC to isolate EVs from cancer patient clinical samples, including plasma, urine, and lavage, demonstrating the device's broad applicability to cancers and other diseases. Finally, the ability of ExoTIC to efficiently isolate EVs from small sample volumes opens up avenues for preclinical studies in small animal tumor models and for point-of-care EV-based clinical testing from fingerprick quantities (10-100 μL) of blood.

  View details for DOI 10.1021/acsnano.7b04878

  View details for PubMedID 29090896

• Deactivated CRISPR Associated Protein 9 for Minor-Allele Enrichment in Cell-Free DNA. Clinical chemistry Aalipour, A. n., Dudley, J. C., Park, S. M., Murty, S. n., Chabon, J. J., Boyle, E. A., Diehn, M. n., Gambhir, S. S. 2017

  Abstract

  Cell-free DNA (cfDNA) diagnostics are emerging as a new paradigm of disease monitoring and therapy management. The clinical utility of these diagnostics is relatively limited by a low signal-to-noise ratio, such as with low allele frequency (AF) mutations in cancer. While enriching for rare alleles to increase their AF before sample analysis is one strategy that can greatly improve detection capability, current methods are limited in their generalizability, ease of use, and applicability to point mutations.Leveraging the robust single-base-pair specificity and generalizability of the CRISPR associated protein 9 (Cas9) system, we developed a deactivated Cas9 (dCas9)-based method of minor-allele enrichment capable of efficient single-target and multiplexed enrichment. The dCas9 protein was complexed with single guide RNAs targeted to mutations of interest and incubated with cfDNA samples containing mutant strands at low abundance. Mutation-bound dCas9 complexes were isolated, dissociated, and the captured DNA purified for downstream use.Targeting the 3 most common epidermal growth factor receptor mutations (exon 19 deletion, T790M, L858R) found in nonsmall-cell lung cancer (NSCLC), we achieved >20-fold increases in AF and detected mutations by use of qPCR at an AF of 0.1%. In a cohort of 18 NSCLC patient-derived cfDNA samples, our method enabled detection of 8 out of 13 mutations that were otherwise undetected by qPCR.The dCas9 method provides important application of the CRISPR/Cas9 system outside the realm of genome editing and can provide a step forward for the detection capability of cfDNA diagnostics.

  View details for PubMedID 29038154

• Engineering Intracellularly Retained Gaussia Luciferase Reporters for Improved Biosensing and Molecular Imaging Applications. ACS chemical biology Gaur, S. n., Bhargava-Shah, A. n., Hori, S. n., Afjei, R. n., Sekar, T. V., Gambhir, S. S., Massoud, T. F., Paulmurugan, R. n. 2017

  Abstract

  Gaussia luciferase (GLUC) is a bioluminescent reporter protein of increasing importance. As a secretory protein, it has increased sensitivity in vitro and in vivo (∼20 000-fold, and ∼1000-fold, respectively) over its competitor, secreted alkaline phosphatase. Unfortunately, this same advantageous secretory nature of GLUC limits its usefulness for many other possible intracellular applications, e.g., imaging signaling pathways in intact cells, in vivo imaging, and in developing molecular imaging biosensors to study protein-protein interactions and protein folding. Hence, to widen the research applications of GLUC, we developed engineered variants that increase its intracellular retention both by modifying the N-terminal secretory signal peptide and by tagging additional sequences to its C-terminal region. We found that when GLUC was expressed in mammalian cells, its N-terminal secretory signal peptide comprising amino acids 1-16 was essential for GLUC folding and functional activity in addition to its inherent secretory property. Modification of the C-terminus of GLUC by tagging a four amino acid (KDEL) endoplasmic reticulum targeting peptide in multiple repeats significantly improved its intracellular retention, with little impact on its folding and enzymatic activity. We used stable cells expressing this engineered GLUC with KDEL repeats to monitor chemically induced endoplasmic reticulum stress on cells. Additionally, we engineered an apoptotic sensor using modified variants of GLUC containing a four amino acid caspase substrate peptide (DEVD) between the GLUC protein and the KDEL repeats. Its use in cell culture resulted in increased GLUC secretion in the growth medium when cells were treated with the chemotherapeutic drugs doxorubicin, paclitaxel, and carboplatin. We thus successfully engineered a new variant GLUC protein that is retained inside cells rather than secreted extracellularly. We validated this novel reporter by incorporating it in biosensors for detection of cellular endoplasmic reticulum stress and caspase activation. This new molecularly engineered enzymatic reporter has the potential for widespread applications in biological research.

  View details for PubMedID 28767220

• Future cancer research priorities in the USA: A Lancet Oncology Commission Future cancer research priorities in the USA: A Lancet Oncology Commission Jaffee, E. M., Dang, C. V., Agus, D. B., Alexaner, B. M., Anderson, K. C., Ashworth, A., Barker, A. D., Bastani, R., Bhatia, S., Bluestone, J. A., Brawley, O., Butte, A. J., Coit, D. G., Davidson, N. E., Davis, M., DePinho, R. A., Diasio, R. B., Draetta, G., Frazier, A. L., Futreal, A., Gambhir, S. S., Ganz, P. A., Garraway, L., Gerson, S., Gupta, S., et al 2017; 18 (11): E653-E706
• A First Report on [18F]FPRGD2 PET/CT Imaging in Multiple Myeloma. Contrast media & molecular imaging Withofs, N. n., Cousin, F. n., De Prijck, B. n., Bonnet, C. n., Hustinx, R. n., Gambhir, S. S., Beguin, Y. n., Caers, J. n. 2017; 2017: 6162845

  Abstract

  An observational study was set up to assess the feasibility of [18F]FPRGD2 PET/CT for imaging patients with multiple myeloma (MM) and to compare its detection rate with low dose CT alone and combined [18F]NaF/[18F]FDG PET/CT images. Four patients (2 newly diagnosed patients and 2 with relapsed MM) were included and underwent whole-body PET/CT after injection of [18F]FPRGD2. The obtained images were compared with results of low dose CT and already available results of a combined [18F]NaF/[18F]FDG PET/CT. In total, 81 focal lesions (FLs) were detected with PET/CT and an underlying bone destruction or fracture was seen in 72 (89%) or 8 (10%) FLs, respectively. Fewer FLs (54%) were detected by [18F]FPRGD2 PET/CT compared to low dose CT (98%) or [18F]NaF/[18F]FDG PET/CT (70%) and all FLs detected with [18F]FPRGD2 PET were associated with an underlying bone lesion. In one newly diagnosed patient, more [18F]FPRGD2 positive lesions were seen than [18F]NaF/[18F]FDG positive lesions. This study suggests that [18F]FPRGD2 PET/CT might be less useful for the detection of myeloma lesions in patients with advanced disease as all FLs with [18F]FPRGD2 uptake were already detected with CT alone.

  View details for PubMedID 29097930

  View details for PubMedCentralID PMC5612716

• Cancer Diagnostics: On-target Probes for Early Detection Nature Biomedical Engineering Hori, S. S., Willemieke, S., Tummers, S., Gambhir, S. S. 2017; 1 (0062): 1-3
• [F-18]GE-180 PET Detects Reduced Microglia Activation After LM11A-31 Therapy in a Mouse Model of Alzheimer's Disease THERANOSTICS James, M. L., Belichenko, N. P., Shuhendler, A. J., Hoehne, A., Andrews, L. E., Condon, C., Nguyen, T. V., Reiser, V., Jones, P., Trigg, W., Rao, J., Gambhir, S. S., Longo, F. M. 2017; 7 (6): 1422-1436

  Abstract

  Microglial activation is a key pathological feature of Alzheimer's disease (AD). PET imaging of translocator protein 18 kDa (TSPO) is a strategy to detect microglial activation in vivo. Here we assessed flutriciclamide ([(18)F]GE-180), a new second-generation TSPO-PET radiotracer, for its ability to monitor response to LM11A-31, a novel AD therapeutic in clinical trials. AD mice displaying pathology were treated orally with LM11A-31 for 3 months. Subsequent [(18)F]GE-180-PET imaging revealed significantly lower signal in cortex and hippocampus of LM11A-31-treated AD mice compared to those treated with vehicle, corresponding with decreased levels of TSPO immunostaining and microglial Iba1 immunostaining. In addition to detecting decreased microglial activation following LM11A-31 treatment, [(18)F]GE-180 identified activated microglia in AD mice with greater sensitivity than another second-generation TSPO radiotracer, [(18)F]PBR06. Together, these data demonstrate the promise of [(18)F]GE-180 as a potentially sensitive tool for tracking neuroinflammation in AD mice and for monitoring therapeutic modulation of microglial activation.

  View details for DOI 10.7150/thno.17666

  View details for PubMedID 28529627

• Withaferin A and its potential role in glioblastoma (GBM) JOURNAL OF NEURO-ONCOLOGY Dhami, J., Chang, E., Gambhir, S. S. 2017; 131 (2): 201-211

  Abstract

  Within the Ayurvedic medical tradition of India, Ashwagandha (Withania somnifera) is a well-known herb. A large number of withanolides have been isolated from both its roots and its leaves and many have been assessed for their pharmacological activities. Amongst them, Withaferin A is one of its most bioactive phytoconstituents. Due to the lactonal steroid's potential to modulate multiple oncogenic pathways, Withaferin A has gained much attention as a possible anti-neoplastic agent. This review focuses on the use of Withaferin A alone, or in combination with other treatments, as a newer option for therapy against the most aggressive variant of brain tumors, Glioblastoma. We survey the various studies that delineate Withaferin A's anticancer mechanisms, its toxicity profiles, its pharmacokinetics and pharmacodynamics and its immuno-modulating properties.

  View details for DOI 10.1007/s11060-016-2303-x

  View details for Web of Science ID 000394342500001

  View details for PubMedID 27837436

• A Pixel-Pitch-Matched Ultrasound Receiver for 3D Photoacoustic Imaging with Integrated Delta-Sigma Beamformer in 28nm UTBB FDSOI Chen, M., Perez, A., Kothapalli, S., Cathelin, P., Cathelin, A., Gambhir, S., Murmann, B., IEEE IEEE. 2017: 456

  View details for Web of Science ID 000403393800192

• Prospective Evaluation of 68Ga-RM2 PET/MRI in Patients with Biochemical Recurrence of Prostate Cancer and Negative Conventional Imaging. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Minamimoto, R. n., Sonni, I. n., Hancock, S. n., Vasanawala, S. n., Loening, A. n., Gambhir, S. S., Iagaru, A. n. 2017

  Abstract

  Purpose:68Ga-labeled DOTA-4-amino-1-carboxymethyl-piperidine-D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 (68Ga-RM2) is a synthetic bombesin receptor antagonist that targets gastrin-releasing peptide receptors (GRPr). GRPr proteins are highly overexpressed in several human tumors, including prostate cancer. We present data from the use of 68Ga-RM2 in patients with biochemical recurrence (BCR) of prostate cancer (PC) and negative conventional imaging (CI). Methods: We enrolled 32 men with BCR PC, 59-83 year-old (mean±standard deviation (SD): 68.7±6.4). Imaging started at 40-69 minutes (mean±SD: 50.5±6.8) after injection of 133.2-151.7 MBq (mean±SD: 140.6±7.4) of 68Ga-RM2 using a time-of-flight (TOF)-enabled simultaneous positron emission tomography (PET) / magnetic resonance imaging (MRI) scanner. T1-weighted (T1w), T2-weighted (T2w) and diffusion-weighted images (DWI) were acquired. Results: All patients had rising prostate specific antigen (PSA) (range: 0.3-119.0 ng/mL; mean±SD: 10.1 ± 21.3) and negative CI (CT or MRI, and 99mTc MDP bone scan) prior to enrollment. The observed 68Ga-RM2 PET detection rate was 71.8%. 68Ga-RM2 PET identified recurrent prostate cancer in 23 of the 32 participants, while the simultaneous MRI scan identified findings compatible with recurrent prostate cancer in 11 of the 32 patients. PSA velocity (PSAv) values were 0.32±0.59 ng/ml/year (range: 0.04-1.9) in patients with negative PET scans and 2.51±2.16 ng/ml/year (range: 0.13-8.68) in patients with positive PET scans (P: 0.006). Conclusion:68Ga-RM2 PET can be used for assessment of GRPr expression in patients with BCR PC. High uptake in multiple areas compatible with cancer lesions suggests that 68Ga-RM2 is a promising PET radiopharmaceutical for localization of disease in participants with BCR PC and negative CI.

  View details for PubMedID 29084827

• Longitudinal Monitoring of Antibody Responses against Tumor Cells Using Magneto-nanosensors with a Nanoliter of Blood. Nano letters Lee, J. R., Chan, C. T., Ruderman, D. n., Chuang, H. Y., Gaster, R. S., Atallah, M. n., Mallick, P. n., Lowe, S. W., Gambhir, S. S., Wang, S. X. 2017; 17 (11): 6644–52

  Abstract

  Each immunoglobulin isotype has unique immune effector functions. The contribution of these functions in the elimination of pathogens and tumors can be determined by monitoring quantitative temporal changes in isotype levels. Here, we developed a novel technique using magneto-nanosensors based on the effect of giant magnetoresistance (GMR) for longitudinal monitoring of total and antigen-specific isotype levels with high precision, using as little as 1 nL of serum. Combining in vitro serologic measurements with in vivo imaging techniques, we investigated the role of the antibody response in the regression of firefly luciferase (FL)-labeled lymphoma cells in spleen, kidney, and lymph nodes in a syngeneic Burkitt's lymphoma mouse model. Regression status was determined by whole body bioluminescent imaging (BLI). The magneto-nanosensors revealed that anti-FL IgG2a and total IgG2a were elevated and sustained in regression mice compared to non-regression mice (p < 0.05). This platform shows promise for monitoring immunotherapy, vaccination, and autoimmunity.

  View details for PubMedID 28990786

• Visualizing Nerve Injury in a Neuropathic Pain Model with [(18)F]FTC-146 PET/MRI. Theranostics Shen, B. n., Behera, D. n., James, M. L., Reyes, S. T., Andrews, L. n., Cipriano, P. W., Klukinov, M. n., Lutz, A. B., Mavlyutov, T. n., Rosenberg, J. n., Ruoho, A. E., McCurdy, C. R., Gambhir, S. S., Yeomans, D. C., Biswal, S. n., Chin, F. T. 2017; 7 (11): 2794–2805

  Abstract

  The ability to locate nerve injury and ensuing neuroinflammation would have tremendous clinical value for improving both the diagnosis and subsequent management of patients suffering from pain, weakness, and other neurologic phenomena associated with peripheral nerve injury. Although several non-invasive techniques exist for assessing the clinical manifestations and morphological aspects of nerve injury, they often fail to provide accurate diagnoses due to limited specificity and/or sensitivity. Herein, we describe a new imaging strategy for visualizing a molecular biomarker of nerve injury/neuroinflammation, i.e., the sigma-1 receptor (S1R), in a rat model of nerve injury and neuropathic pain. The two-fold higher increase of S1Rs was shown in the injured compared to the uninjured nerve by Western blotting analyses. With our novel S1R-selective radioligand, [(18)F]FTC-146 (6-(3-[(18)F]fluoropropyl)-3-(2-(azepan-1-yl)ethyl)benzo[d]thiazol-2(3H)-one), and positron emission tomography-magnetic resonance imaging (PET/MRI), we could accurately locate the site of nerve injury created in the rat model. We verified the accuracy of this technique by ex vivo autoradiography and immunostaining, which demonstrated a strong correlation between accumulation of [(18)F]FTC-146 and S1R staining. Finally, pain relief could also be achieved by blocking S1Rs in the neuroma with local administration of non-radioactive [(19)F]FTC-146. In summary, [(18)F]FTC-146 S1R PET/MR imaging has the potential to impact how we diagnose, manage and treat patients with nerve injury, and thus warrants further investigation.

  View details for PubMedID 28824716

• Molecular profiling of single circulating tumor cells from lung cancer patients PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Park, S., Wong, D. J., Ooi, C. C., Kurtz, D. M., Vermesh, O., Aalipour, A., Suh, S., Pian, K. L., Chabon, J. J., Lee, S. H., Jamali, M., Say, C., Carter, J. N., Lee, L. P., Kuschner, W. G., Schwartz, E. J., Shrager, J. B., Neal, J. W., Wakelee, H. A., Diehn, M., Nair, V. S., Wang, S. X., Gambhir, S. S. 2016; 113 (52): E8379-E8386

  Abstract

  Circulating tumor cells (CTCs) are established cancer biomarkers for the "liquid biopsy" of tumors. Molecular analysis of single CTCs, which recapitulate primary and metastatic tumor biology, remains challenging because current platforms have limited throughput, are expensive, and are not easily translatable to the clinic. Here, we report a massively parallel, multigene-profiling nanoplatform to compartmentalize and analyze hundreds of single CTCs. After high-efficiency magnetic collection of CTC from blood, a single-cell nanowell array performs CTC mutation profiling using modular gene panels. Using this approach, we demonstrated multigene expression profiling of individual CTCs from non-small-cell lung cancer (NSCLC) patients with remarkable sensitivity. Thus, we report a high-throughput, multiplexed strategy for single-cell mutation profiling of individual lung cancer CTCs toward minimally invasive cancer therapy prediction and disease monitoring.

  View details for DOI 10.1073/pnas.1608461113

  View details for PubMedID 27956614

• Practical ImmunoPET radiotracer design considerations for human immune checkpoint imaging. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Mayer, A. T., Natarajan, A., Gordon, S., Maute, R., McCracken, M., Ring, A., Weissman, I., Gambhir, S. S. 2016

  Abstract

  Immune checkpoint blockade has emerged as a promising cancer treatment paradigm. Unfortunately, there are still a large number of patients and malignancies that do not respond to therapy. A major barrier to validating biomarkers for the prediction and monitoring of responders to clinical checkpoint blockade has been the lack of imaging tools to accurately assess dynamic immune checkpoint expression. Here, we sought to optimize noninvasive immuno-PET imaging of human programmed death-ligand 1 (PD-L1) expression, in a preclinical model, using a small high-affinity engineered protein scaffold (HAC-PD1). Six HAC-PD1 radiotracer variants were developed and used in preclinical imaging and biodistribution studies to assess their ability to detect human PD-L1 expression in vivo. Radiotracer design modifications included chelate, glycosylation, and radiometal. HACA-PD1 was adopted as the naming convention for aglycosylated tracer variants. NOD scid γ-(NSG) mice were inoculated with subcutaneous tumors engineered to either be constitutively positive (CT26 hPD-L1) or be negative (ΔmPD-L1 CT26) for human PD-L1 expression. When the tumors had grown to an average size of 1 cm in diameter, mice were injected with 0.75-2.25 MBq (∼10 μg) of an engineered radiotracer variant and imaged. At 1 h after injection, organs were harvested for biodistribution. Of the practical immuno-PET tracer modifications considered, glycosylation was the most prominent design factor affecting tracer uptake, specificity, and clearance. In imaging studies, aglycosylated (64)Cu-NOTA-HACA-PD1 most accurately visualized human PD-L1 expression in vivo. We reasoned that because of the scaffold's small size (14 kDa), its pharmacokinetics may be suitable for labeling with the short-lived and widely clinically available radiometal (68)Ga. At 1 h after injection, (68)Ga-NOTA-HACA-PD1 and (68)Ga-DOTA-HACA-PD1 exhibited promising target-to-background ratios in ex vivo biodistribution studies (12.3 and 15.2 tumor-to-muscle ratios, respectively). Notably, all HAC-PD1 radiotracer variants enabled much earlier detection of human PD-L1 expression (1 h after injection) than previously reported radiolabeled antibodies (>24 h after injection). This work provides a template for assessing immuno-PET tracer design parameters and supports the translation of small engineered protein radiotracers for imaging human immune checkpoints.

  View details for PubMedID 27980047

• A Clinical Wide-Field Fluorescence Endoscopic Device for Molecular Imaging Demonstrating Cathepsin Protease Activity in Colon Cancer MOLECULAR IMAGING AND BIOLOGY Sensarn, S., Zavaleta, C. L., Segal, E., Rogalla, S., Lee, W., Gambhir, S. S., Bogyo, M., Contag, C. H. 2016; 18 (6): 820–29

  View details for DOI 10.1007/s11307-016-0956-7

  View details for Web of Science ID 000387367100004

• Can multispectral optoacoustic tomography replace sentinel lymph biopsy in melanoma? Annals of translational medicine Leong, S. P., Kothapalli, S., Gambhir, S. S. 2016; 4 (24): 517-?

  View details for DOI 10.21037/atm.2016.12.31

  View details for PubMedID 28149879

  View details for PubMedCentralID PMC5233521

• A Clinical Wide-Field Fluorescence Endoscopic Device for Molecular Imaging Demonstrating Cathepsin Protease Activity in Colon Cancer. Molecular imaging and biology Sensarn, S., Zavaleta, C. L., Segal, E., Rogalla, S., Lee, W., Gambhir, S. S., Bogyo, M., Contag, C. H. 2016; 18 (6): 820-829

  Abstract

  Early and effective detection of cancers of the gastrointestinal tract will require novel molecular probes and advances in instrumentation that can reveal functional changes in dysplastic and malignant tissues. Here, we describe adaptation of a wide-field clinical fiberscope to perform wide-field fluorescence imaging while preserving its white-light capability for the purpose of providing wide-field fluorescence imaging capability to point-of-care microscopes.We developed and used a fluorescent fiberscope to detect signals from a quenched probe, BMV109, that becomes fluorescent when cleaved by, and covalently bound to, active cathepsin proteases. Cathepsins are expressed in inflammation- and tumor-associated macrophages as well as directly from tumor cells and are a promising target for cancer imaging. The fiberscope has a 1-mm outer diameter enabling validation via endoscopic exams in mice, and therefore we evaluated topically applied BMV109 for the ability to detect colon polyps in an azoxymethane-induced colon tumor model in mice.This wide-field endoscopic imaging device revealed consistent and clear fluorescence signals from BMV109 that specifically localized to the polypoid regions as opposed to the normal adjacent colon tissue (p < 0.004) in the murine colon carcinoma model.The sensitivity of detection of BMV109 with the fluorescence fiberscope suggested utility of these tools for early detection at hard-to-reach sites. The fiberscope was designed to be used in conjunction with miniature, endoscope-compatible fluorescence microscopes for dual wide-field and microscopic cancer detection.

  View details for PubMedID 27154508

• Clinically Approved Nanoparticle Imaging Agents JOURNAL OF NUCLEAR MEDICINE Thakor, A. S., Jokerst, J. V., Ghanouni, P., Campbell, J. L., Mittra, E., Gambhir, S. S. 2016; 57 (12): 1833–37

  View details for DOI 10.2967/jnumed.116.181362

  View details for Web of Science ID 000389106800005

• Clinically Approved Nanoparticle Imaging Agents. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Thakor, A. S., Jokerst, J. V., Ghanouni, P., Campbell, J., Mittra, E., Gambhir, S. S. 2016

  Abstract

  Nanoparticles are a new class of imaging agent used for both anatomic and molecular imaging. Nanoparticle-based imaging exploits the signal intensity, stability, and biodistribution behavior of submicron-diameter molecular imaging agents. This review focuses on nanoparticles used in human medical imaging, with an emphasis on radionuclide imaging and MRI. Newer nanoparticle platforms are also discussed in relation to theranostic and multimodal uses.

  View details for PubMedID 27738007

• A transgenic mouse model expressing an ER alpha folding biosensor reveals the effects of Bisphenol A on estrogen receptor signaling SCIENTIFIC REPORTS Sekar, T. V., Foygel, K., Massoud, T. F., Gambhir, S. S., Paulmurugan, R. 2016; 6

  Abstract

  Estrogen receptor-α (ERα) plays an important role in normal and abnormal physiology of the human reproductive system by interacting with the endogenous ligand estradiol (E2). However, other ligands, either analogous or dissimilar to E2, also bind to ERα. This may create unintentional activation of ER signaling in reproductive tissues that can lead to cancer development. We developed a transgenic mouse model that constitutively expresses a firefly luciferase (FLuc) split reporter complementation biosensor (NFLuc-ER-LBDG521T-CFLuc) to simultaneously evaluate the dynamics and potency of ligands that bind to ERα. We first validated this model using various ER ligands, including Raloxifene, Diethylstilbestrol, E2, and 4-hydroxytamoxifen, by employing FLuc-based optical bioluminescence imaging of living mice. We then used the model to investigate the carcinogenic property of Bisphenol A (BPA), an environmental estrogen, by long-term exposure at full and half environmental doses. We showed significant carcinogenic effects on female animals while revealing activated downstream ER signaling as measured by bioluminescence imaging. BPA induced tumor-like outgrowths in female transgenic mice, histopathologically confirmed to be neoplastic and epithelial in origin. This transgenic mouse model expressing an ERα folding-biosensor is useful in evaluation of estrogenic ligands and their downstream effects, and in studying environmental estrogen induced carcinogenesis in vivo.

  View details for DOI 10.1038/srep34788

  View details for PubMedID 27721470

• A Cystine Knot Peptide Targeting Integrin alpha(v)beta(6) for Photoacoustic and Fluorescence Imaging of Tumors in Living Subjects JOURNAL OF NUCLEAR MEDICINE Zhang, C., Kimura, R., Abou-Elkacem, L., Levi, J., Xu, L., Gambhir, S. S. 2016; 57 (10): 1629-1634

  Abstract

  Photoacoustic imaging is a nonionizing biomedical imaging modality with higher resolution and imaging depth than fluorescence imaging, which has greater sensitivity. The combination of the 2 imaging modalities could improve the detection of cancer. Integrin αvβ6 is a cell surface marker overexpressed in many different cancers. Here, we report the development and evaluation of a dye-labeled cystine knot peptide, which selectively recognizes integrin αvβ6 with high affinity, for photoacoustic and fluorescence imaging. The new dual-modality probe may find clinical application in cancer diagnosis and intraoperative imaging of integrin αvβ6-positive tumors.An engineered cystine knot peptide, R01, that recognizes integrin αvβ6 was labeled with Atto 740 (A740) and evaluated for its specific cell uptake and its sensitivity threshold. A740-R01 was injected via the tail vein into nude mice xenografted with A431 (integrin αvβ6-positive) or 293T (integrin αvβ6-negative) tumors. Photoacoustic and fluorescence scans of tumors were acquired before and at 0.5, 1, 2, and 4 h after injection of A740-R01. Dynamic photoacoustic scans of various normal organs were also acquired. Ex vivo fluorescence imaging of tissues was performed 1 h after injection.The A740-R01 demonstrated integrin αvβ6-dependent binding to A431 cells in culture. Sensitivity studies indicated that the probe may potentially detect lesions as small as 1 or 6 mm(3) by fluorescence or photoacoustic imaging, respectively. The photoacoustic and fluorescence signals of A431 xenografts at 1 h after injection were 1.87 ± 0.25 arbitrary units (AU) and 8.27 ± 0.87 AU, respectively. Target specificity was confirmed by low tumor uptake in 293T tumors at 1 h after injection (1.07 ± 0.15 AU and 1.10 ± 0.14 AU for photoacoustic and fluorescence signals, respectively). A740-R01 exhibited hepatobiliary clearance marked by high uptake in the liver, spleen, and intestine but low uptake in the kidneys.A740-R01 specifically targeted integrin αvβ6 with low nanomolar affinity. A740-R01 was able to detect integrin αvβ6 both in vitro and in vivo by photoacoustic and fluorescence imaging. A740-R01 is able to detect αvβ6-positive tumors in living subjects and may have clinical application in cancer diagnosis and real-time image-guided surgery.

  View details for DOI 10.2967/jnumed.115.169383

  View details for Web of Science ID 000384961900031

  View details for PubMedID 27230926

• Quantitative photoacoustic image reconstruction improves accuracy in deep tissue structures. Biomedical optics express Mastanduno, M. A., Gambhir, S. S. 2016; 7 (10): 3811-3825

  Abstract

  Photoacoustic imaging (PAI) is emerging as a potentially powerful imaging tool with multiple applications. Image reconstruction for PAI has been relatively limited because of limited or no modeling of light delivery to deep tissues. This work demonstrates a numerical approach to quantitative photoacoustic image reconstruction that minimizes depth and spectrally derived artifacts. We present the first time-domain quantitative photoacoustic image reconstruction algorithm that models optical sources through acoustic data to create quantitative images of absorption coefficients. We demonstrate quantitative accuracy of less than 5% error in large 3 cm diameter 2D geometries with multiple targets and within 22% error in the largest size quantitative photoacoustic studies to date (6cm diameter). We extend the algorithm to spectral data, reconstructing 6 varying chromophores to within 17% of the true values. This quantitiative PA tomography method was able to improve considerably on filtered-back projection from the standpoint of image quality, absolute, and relative quantification in all our simulation geometries. We characterize the effects of time step size, initial guess, and source configuration on final accuracy. This work could help to generate accurate quantitative images from both endogenous absorbers and exogenous photoacoustic dyes in both preclinical and clinical work, thereby increasing the information content obtained especially from deep-tissue photoacoustic imaging studies.

  View details for DOI 10.1364/BOE.7.003811

  View details for PubMedID 27867695

  View details for PubMedCentralID PMC5102520

• Quantitative photoacoustic image reconstruction improves accuracy in deep tissue structures BIOMEDICAL OPTICS EXPRESS Mastanduno, M. A., Gambhir, S. S. 2016; 7 (10): 3811-3825

  View details for DOI 10.1364/BOE.7.003811

  View details for Web of Science ID 000385418200001

• Imaging approaches to optimize molecular therapies SCIENCE TRANSLATIONAL MEDICINE Weissleder, R., Schwaiger, M. C., Gambhir, S. S., Hricak, H. 2016; 8 (355)

  Abstract

  Imaging, including its use for innovative tissue sampling, is slowly being recognized as playing a pivotal role in drug development, clinical trial design, and more effective delivery and monitoring of molecular therapies. The challenge is that, while a considerable number of new imaging technologies and new targeted tracers have been developed for cancer imaging in recent years, the technologies are neither evenly distributed nor evenly implemented. Furthermore, many imaging innovations are not validated and are not ready for widespread use in drug development or in clinical trial designs. Inconsistent and often erroneous use of terminology related to quantitative imaging biomarkers has also played a role in slowing their development and implementation. We examine opportunities for, and challenges of, the use of imaging biomarkers to facilitate development of molecular therapies and to accelerate progress in clinical trial design. In the future, in vivo molecular imaging, image-guided tissue sampling for mutational analyses ("high-content biopsies"), and noninvasive in vitro tests ("liquid biopsies") will likely be used in various combinations to provide the best possible monitoring and individualized treatment plans for cancer patients.

  View details for DOI 10.1126/scitranslmed.aaf3936

  View details for Web of Science ID 000384015200001

  View details for PubMedID 27605550

• Multimodality Molecular Imaging of Cardiac Cell Transplantation: Part II. In Vivo Imaging of Bone Marrow Stromal Cells in Swine with PET/CT and MR Imaging. Radiology Parashurama, N., Ahn, B., Ziv, K., Ito, K., Paulmurugan, R., Willmann, J. K., Chung, J., Ikeno, F., Swanson, J. C., Merk, D. R., Lyons, J. K., Yerushalmi, D., Teramoto, T., Kosuge, H., Dao, C. N., Ray, P., Patel, M., Chang, Y., Mahmoudi, M., Cohen, J. E., Goldstone, A. B., Habte, F., Bhaumik, S., Yaghoubi, S., Robbins, R. C., Dash, R., Yang, P. C., Brinton, T. J., Yock, P. G., McConnell, M. V., Gambhir, S. S. 2016; 280 (3): 826-836

  Abstract

  Purpose To quantitatively determine the limit of detection of marrow stromal cells (MSC) after cardiac cell therapy (CCT) in swine by using clinical positron emission tomography (PET) reporter gene imaging and magnetic resonance (MR) imaging with cell prelabeling. Materials and Methods Animal studies were approved by the institutional administrative panel on laboratory animal care. Seven swine received 23 intracardiac cell injections that contained control MSC and cell mixtures of MSC expressing a multimodality triple fusion (TF) reporter gene (MSC-TF) and bearing superparamagnetic iron oxide nanoparticles (NP) (MSC-TF-NP) or NP alone. Clinical MR imaging and PET reporter gene molecular imaging were performed after intravenous injection of the radiotracer fluorine 18-radiolabeled 9-[4-fluoro-3-(hydroxyl methyl) butyl] guanine ((18)F-FHBG). Linear regression analysis of both MR imaging and PET data and nonlinear regression analysis of PET data were performed, accounting for multiple injections per animal. Results MR imaging showed a positive correlation between MSC-TF-NP cell number and dephasing (dark) signal (R(2) = 0.72, P = .0001) and a lower detection limit of at least approximately 1.5 × 10(7) cells. PET reporter gene imaging demonstrated a significant positive correlation between MSC-TF and target-to-background ratio with the linear model (R(2) = 0.88, P = .0001, root mean square error = 0.523) and the nonlinear model (R(2) = 0.99, P = .0001, root mean square error = 0.273) and a lower detection limit of 2.5 × 10(8) cells. Conclusion The authors quantitatively determined the limit of detection of MSC after CCT in swine by using clinical PET reporter gene imaging and clinical MR imaging with cell prelabeling. (©) RSNA, 2016 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.2016151150

  View details for PubMedID 27332865

• Multimodality Molecular Imaging of Cardiac Cell Transplantation: Part I. Reporter Gene Design, Characterization, and Optical in Vivo Imaging of Bone Marrow Stromal Cells after Myocardial Infarction. Radiology Parashurama, N., Ahn, B., Ziv, K., Ito, K., Paulmurugan, R., Willmann, J. K., Chung, J., Ikeno, F., Swanson, J. C., Merk, D. R., Lyons, J. K., Yerushalmi, D., Teramoto, T., Kosuge, H., Dao, C. N., Ray, P., Patel, M., Chang, Y., Mahmoudi, M., Cohen, J. E., Goldstone, A. B., Habte, F., Bhaumik, S., Yaghoubi, S., Robbins, R. C., Dash, R., Yang, P. C., Brinton, T. J., Yock, P. G., McConnell, M. V., Gambhir, S. S. 2016; 280 (3): 815-825

  Abstract

  Purpose To use multimodality reporter-gene imaging to assess the serial survival of marrow stromal cells (MSC) after therapy for myocardial infarction (MI) and to determine if the requisite preclinical imaging end point was met prior to a follow-up large-animal MSC imaging study. Materials and Methods Animal studies were approved by the Institutional Administrative Panel on Laboratory Animal Care. Mice (n = 19) that had experienced MI were injected with bone marrow-derived MSC that expressed a multimodality triple fusion (TF) reporter gene. The TF reporter gene (fluc2-egfp-sr39ttk) consisted of a human promoter, ubiquitin, driving firefly luciferase 2 (fluc2), enhanced green fluorescent protein (egfp), and the sr39tk positron emission tomography reporter gene. Serial bioluminescence imaging of MSC-TF and ex vivo luciferase assays were performed. Correlations were analyzed with the Pearson product-moment correlation, and serial imaging results were analyzed with a mixed-effects regression model. Results Analysis of the MSC-TF after cardiac cell therapy showed significantly lower signal on days 8 and 14 than on day 2 (P = .011 and P = .001, respectively). MSC-TF with MI demonstrated significantly higher signal than MSC-TF without MI at days 4, 8, and 14 (P = .016). Ex vivo luciferase activity assay confirmed the presence of MSC-TF on days 8 and 14 after MI. Conclusion Multimodality reporter-gene imaging was successfully used to assess serial MSC survival after therapy for MI, and it was determined that the requisite preclinical imaging end point, 14 days of MSC survival, was met prior to a follow-up large-animal MSC study. (©) RSNA, 2016 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.2016140049

  View details for PubMedID 27308957

• Glioblastoma Multiforme Recurrence: An Exploratory Study of (18)F FPPRGD2 PET/CT. Radiology Iagaru, A., Mosci, C., Mittra, E., Zaharchuk, G., Fischbein, N., Harsh, G., Li, G., Nagpal, S., Recht, L., Gambhir, S. S. 2016; 280 (1): 328-?

  View details for DOI 10.1148/radiol.2016164020

  View details for PubMedID 27322985

• Glioblastoma Multiforme Recurrence Response RADIOLOGY Iagaru, A., Gambhir, S. 2016; 280 (1): 326–27

  View details for Web of Science ID 000378721900042

• Protein biomarkers on tissue as imaged via MALDI mass spectrometry: A systematic approach to study the limits of detection PROTEOMICS van de Ven, S. M., Bemis, K. D., Lau, K., Adusumilli, R., Kota, U., Stolowitz, M., Vitek, O., Mallick, P., Gambhir, S. S. 2016; 16 (11-12): 1660-1669

  Abstract

  MALDI mass spectrometry imaging (MSI) is emerging as a tool for protein and peptide imaging across tissue sections. Despite extensive study, there does not yet exist a baseline study evaluating the potential capabilities for this technique to detect diverse proteins in tissue sections. In this study, we developed a systematic approach for characterizing MALDI-MSI workflows in terms of limits of detection, coefficients of variation, spatial resolution, and the identification of endogenous tissue proteins. Our goal was to quantify these figures of merit for a number of different proteins and peptides, in order to gain more insight in the feasibility of protein biomarker discovery efforts using this technique. Control proteins and peptides were deposited in serial dilutions on thinly sectioned mouse xenograft tissue. Using our experimental setup, coefficients of variation were <30% on tissue sections and spatial resolution was 200 μm (or greater). Limits of detection for proteins and peptides on tissue were in the micromolar to millimolar range. Protein identification was only possible for proteins present in high abundance in the tissue. These results provide a baseline for the application of MALDI-MSI towards the discovery of new candidate biomarkers and a new benchmarking strategy that can be used for comparing diverse MALDI-MSI workflows.

  View details for DOI 10.1002/pmic.201500515

  View details for Web of Science ID 000379049100008

  View details for PubMedID 26970438

• Characterization of Physiologic F-18 FSPG Uptake in Healthy Volunteers RADIOLOGY Mosci, C., Kumar, M., Smolarz, K., Koglin, N., Stephens, A. W., Schwaiger, M., Gambhir, S. S., Mittra, E. S. 2016; 279 (3): 898-905

  Abstract

  Purpose To evaluate the normal biodistribution and kinetics of (S)-4-(3-[18F]fluoropropyl)-l-glutamic acid ((18)F FSPG) in healthy volunteers and to compare (18)F FSPG mean and maximum standardized uptake values (SUVmean and SUVmax, respectively) with those of (18)F fluorodeoxyglucose (FDG) across a variety of organs. Materials and Methods This protocol was reviewed and approved by all appropriate regulatory authorities. An 8-mCi (±10%) dose of (18)F FSPG was given to five subjects (three women, two men), and seven whole-body positron emission tomography (PET) scans were performed 5, 10, 20, 30, 45, 150, and 240 minutes after injection. Regions of interest were analyzed on the resultant (18)F FSPG images to evaluate the kinetics of this radiotracer. The images obtained 45 minutes after injection were used to measure SUVmean and SUVmax in additional regions of the body. These values were compared with similar values obtained with (18)F FDG PET published previously. Descriptive statistics, including average and standard deviation across the five subjects, were used. (18)F FSPG SUVmean and SUVmax were compared. Results On the (18)F FSPG images obtained 45 minutes after injection, there was only low-grade background activity in the majority of analyzed regions. Prominent activity was seen throughout the pancreas. Clearance of the radiotracer through the kidneys and collection in the bladder also were seen. SUV quantification shows notable differences between (18)F FSPG and (18)F FDG in the pancreas ((18)F FSPG SUVmean, 8.2; (18)F FDG SUVmean, 1.3), stomach ((18)F FSPG SUVmax, 3.6; (18)F FDG SUVmax, 1.6), and brain ((18)F FSPG SUVmean, 0.08; (18)F FDG SUVmean, 7.8). The kinetic data showed rapid clearance of the radiotracer from the blood pool and most organs, except the pancreas. Conclusion (18)F FSPG is a PET radiopharmaceutical characterized by rapid clearance from most healthy tissues, except the pancreas and kidneys. A consistent biodistribution pattern was observed with low background uptake. The physiologic uptake of this new radiotracer throughout the body is described in more detail, which is important for improved interpretative accuracy and understanding potential clinical applications. (©) RSNA, 2016.

  View details for DOI 10.1148/radiol.2015142000

  View details for Web of Science ID 000378719700028

  View details for PubMedID 26785040

• Pilot prospective evaluation of F-18-FPPRGD(2) PET/CT in patients with cervical and ovarian cancer EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Minamimoto, R., Karam, A., Jamali, M., Barkhodari, A., Gambhir, S. S., Dorigo, O., Iagaru, A. 2016; 43 (6): 1047-1055

  Abstract

  We report the effect of antiangiogenic therapy on the biodistribution of (18)F-FPPRGD2 (a surrogate biomarker of integrin αvβ3 expression), and the potential of (18)F-FPPRGD2 to predict the prognosis in patients with cervical cancer and ovarian cancer in this clinical scenario.Data from six women, age range 30 - 59 years (mean ± SD 44.0 ± 12.5 years), who had undergone a (18)F-FPPRGD2 PET/CT scan and bevacizumab-containing therapy were prospectively collected and analyzed. We compared baseline (18)F-FPPRGD2 and (18)F-FDG uptake in the lesions and tumor-to-background (T/B) ratios. The maximum and mean (18)F-FPPRGD2 standardized uptake values (SUVmax and SUVmean) were recorded for 13 normal organs, as well as in all the identified malignant lesions on the pretreatment scan and the 1-week post-treatment scan. We also measured changes in (18)F-FPPRGD2 uptake from before to 1 week after treatment, and compared them to the changes in (18)F-FDG uptake from before to 6 weeks after treatment. Treatment outcomes were correlated with these changes.The uptake in lesions and T/B ratio of (18)F-FPPRGD2 were lower than those of (18)F-FDG (SUVmax 3.7 ± 1.3 vs. 6.0 ± 1.8, P < 0.001; SUVmean 2.6 ± 0.7 vs. 4.2 ± 1.3, P < 0.001; T/B ratio based on SUVmax 2.4 ± 1.0 vs. 2.6 ± 1.0, P < 0.04; T/B ratio based on SUVmean 1.9 ± 0.6 vs. 2.4 ± 1.0, P < 0.003). One patient did not return for the follow-up scan and in another patient no lesions were identified on the pretreatment scan. (18)F-FPPRGD2 uptake in lesions in the remaining four patients had significantly changed 1 week after treatment (SUVmean 3.3 ± 1.0 vs. 2.7 ± 1.0, P < 0.001), while uptake in all normal tissues analyzed was not affected by treatment. One patient with clinical disease progression had a decrease in lesional (18)F-FPPRGD2 SUVmean of 1.6 % and in (18)F-FDG SUVmean of 9.4 %. Two patients with a clinical complete response to treatment had decreases in lesional (18)F-FPPRGD2 SUVmean of 25.2 % and 25.0 % and in (18)F-FDG SUVmean of 6.1 % and 71.8 %. One patient with a clinical partial response had a decrease in lesional (18)F-FPPRGD2 SUVmean of 7.9 % and in (18)F-FDG SUVmean of 76.4 %.This pilot study showed that (18)F-FPPRGD2 and (18)F-FDG provide independent information about the biology of ovarian and cervical cancers. Bevacizumab-containing therapy does not affect (18)F-FPPRGD2 uptake in normal organs, but does result in statistically significant changes in lesions. In addition, (18)F-FPPRGD2 may have potential for early prediction of response to such treatments. These preliminary findings have to be confirmed in larger studies.

  View details for DOI 10.1007/s00259-015-3263-7

  View details for Web of Science ID 000374972900008

  View details for PubMedID 26611425

• Alk5 inhibition increases delivery of macromolecular and protein-bound contrast agents to tumors. JCI insight Daldrup-Link, H. E., Mohanty, S., Ansari, C., Lenkov, O., Shaw, A., Ito, K., Hong, S. H., Hoffmann, M., Pisani, L., Boudreau, N., Gambhir, S. S., Coussens, L. M. 2016; 1 (6)

  Abstract

  Limited transendothelial permeability across tumor microvessels represents a significant bottleneck in the development of tumor-specific diagnostic agents and theranostic drugs. Here, we show an approach to increase transendothelial permeability of macromolecular and nanoparticle-based contrast agents via inhibition of the type I TGF-β receptor, activin-like kinase 5 (Alk5), in tumors. Alk5 inhibition significantly increased tumor contrast agent delivery and enhancement on imaging studies, while healthy organs remained relatively unaffected. Imaging data correlated with significantly decreased tumor interstitial fluid pressure, while tumor vascular density remained unchanged. This immediately clinically translatable concept involving Alk5 inhibitor pretreatment prior to an imaging study could be leveraged for improved tumor delivery of macromolecular and nanoparticle-based imaging probes and, thereby, facilitate development of more sensitive imaging tests for cancer diagnosis, enhanced tumor characterization, and personalized, image-guided therapies.

  View details for PubMedID 27182558

• Alk5 inhibition increases delivery of macromolecular and protein-bound contrast agents to tumors JCI INSIGHT Daldrup-Link, H. E., Mohanty, S., Ansari, C., Lenkov, O., Shaw, A., Ito, K., Hong, S., Hoffmann, M., Pisani, L., Boudreau, N., Gambhir, S., Coussens, L. M. 2016; 1 (6)

  View details for DOI 10.1172/jci.insight.85608

  View details for Web of Science ID 000387111300001

• Imaging of tumor-associated system xC-activity with 18F-fluoropropylglutamate (18F-FSPG) PET/CT for intracranial malignancies Sonni, I., Minamimoto, R., Jamali, M., Hatami, N., Berndt, M., Koglin, N., Stephens, A., Iagaru, A., Chin, F., Gambhir, S., Mittra, E. SOC NUCLEAR MEDICINE INC. 2016

  View details for Web of Science ID 000442211000182

• Pilot Comparison of Ga-68-RM2 PET and Ga-68-PSMA-11 PET in Patients with Biochemically Recurrent Prostate Cancer JOURNAL OF NUCLEAR MEDICINE Minamimoto, R., Hancock, S., Schneider, B., Chin, F. T., Jamali, M., Loening, A., Vasanawala, S., Gambhir, S. S., Iagaru, A. 2016; 57 (4): 557-562

  Abstract

  Glu-NH-CO-NH-Lys-(Ahx)-[(68)Ga(HBED-CC)] ((68)Ga-PSMA-11) is a PET tracer that can detect prostate cancer relapses and metastases by binding to the extracellular domain of PSMA.(68)Ga-labeled DOTA-4-amino-1-carboxymethyl-piperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 ((68)Ga-RM2) is a synthetic bombesin receptor antagonist that targets gastrin-releasing peptide receptors. We present pilot data on the biodistribution of these PET tracers in a small cohort of patients with biochemically recurrent prostate cancer.Seven men (mean age ± SD, 74.3 ± 5.9 y) with biochemically recurrent prostate cancer underwent both(68)Ga-PSMA-11 PET/CT and(68)Ga-RM2 PET/MRI scans. SUVmaxand SUVmeanwere recorded for normal tissues and areas of uptake outside the expected physiologic biodistribution.All patients had a rising level of prostate-specific antigen (mean ± SD, 13.5 ± 11.5) and noncontributory results on conventional imaging.(68)Ga-PSMA-11 had the highest physiologic uptake in the salivary glands and small bowel, with hepatobiliary and renal clearance noted, whereas(68)Ga-RM2 had the highest physiologic uptake in the pancreas, with renal clearance noted. Uptake outside the expected physiologic biodistribution did not significantly differ between(68)Ga-PSMA-11 and(68)Ga-RM2; however,(68)Ga-PSMA-11 localized in a lymph node and seminal vesicle in a patient with no abnormal(68)Ga-RM2 uptake. Abdominal periaortic lymph nodes were more easily visualized by(68)Ga-RM2 in two patients because of lack of interference by radioactivity in the small intestine.(68)Ga-PSMA-11 and(68)Ga-RM2 had distinct biodistributions in this small cohort of patients with biochemically recurrent prostate cancer. Additional work is needed to understand the expression of PSMA and gastrin-releasing peptide receptors in different types of prostate cancer.

  View details for DOI 10.2967/jnumed.115.168393

  View details for PubMedID 26659347

• [F-18]FPRGD(2) PET/CT imaging of integrin alpha(v)beta(3) levels in patients with locally advanced rectal carcinoma EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Withofs, N., Martinive, P., Vanderick, J., Bletard, N., Scagnol, I., Mievis, F., Giacomelli, F., Coucke, P., Delvenne, P., Cataldo, D., Gambhir, S. S., Hustinx, R. 2016; 43 (4): 654-662

  View details for DOI 10.1007/s00259-015-3219-y

  View details for Web of Science ID 000371074100007

• [(18)F]FPRGD2 PET/CT imaging of integrin avß3 levels in patients with locally advanced rectal carcinoma. European journal of nuclear medicine and molecular imaging Withofs, N., Martinive, P., Vanderick, J., Bletard, N., Scagnol, I., Mievis, F., Giacomelli, F., Coucke, P., Delvenne, P., Cataldo, D., Gambhir, S. S., Hustinx, R. 2016; 43 (4): 654-662

  Abstract

  Our primary objective was to determine if [(18)F]FPRGD2 PET/CT performed at baseline and/or after chemoradiotherapy (CRT) could predict tumour regression grade (TRG) in locally advanced rectal cancer (LARC). Secondary objectives were to compare baseline [(18)F]FPRGD2 and [(18)F]FDG uptake, to evaluate the correlation between posttreatment [(18)F]FPRGD2 uptake and tumour microvessel density (MVD) and to determine if [(18)F]FPRGD2 and FDG PET/CT could predict disease-free survival.Baseline [(18)F]FPRGD2 and FDG PET/CT were performed in 32 consecutive patients (23 men, 9 women; mean age 63 ± 8 years) with LARC before starting any therapy. A posttreatment [(18)F]FPRGD2 PET/CT scan was performed in 24 patients after the end of CRT (median interval 7 weeks, range 3 - 15 weeks) and before surgery (median interval 4 days, range 1 - 15 days).All LARC showed uptake of both [(18)F]FPRGD2 (SUVmax 5.4 ± 1.5, range 2.7 - 9) and FDG (SUVmax 16.5 ± 8, range 7.1 - 36.5). There was a moderate positive correlation between [(18)F]FPRGD2 and FDG SUVmax (Pearson's r = 0.49, p = 0.0026). There was a moderate negative correlation between baseline [(18)F]FPRGD2 SUVmax and the TRG (Spearman's r = -0.37, p = 0.037), and a [(18)F]FPRGD2 SUVmax of >5.6 identified all patients with a complete response (TRG 0; AUC 0.84, 95 % CI 0.68 - 1, p = 0.029). In the 24 patients who underwent a posttreatment [(18)F]FPRGD2 PET/CT scan the response index, calculated as [(SUVmax1 - SUVmax2)/SUVmax1] × 100 %, was not associated with TRG. Post-treatment [(18)F]FPRGD2 uptake was not correlated with tumour MVD. Neither [(18)F]FPRGD2 nor FDG uptake predicted disease-free survival.Baseline [(18)F]FPRGD2 uptake was correlated with the pathological response in patients with LARC treated with CRT. However, the specificity was too low to consider its clinical routine use.

  View details for DOI 10.1007/s00259-015-3219-y

  View details for PubMedID 26490751

• Comparison of Deconvolution Filters for Photoacoustic Tomography PLOS ONE de Sompel, D. V., Sasportas, L. S., Jokerst, J. V., Gambhir, S. S. 2016; 11 (3)

  View details for DOI 10.1371/journal.pone.0152597

  View details for Web of Science ID 000373121800099

• Mesoporous silica nanoparticles for ultrasound/magnetic resonance imaging and therapeutic drug delivery for stem cell therapy Kempen, P., Campbell, J., Greasley, S., Jones, J., Gambhir, S., Sinclair, R., Jokerst, J. AMER CHEMICAL SOC. 2016

  View details for Web of Science ID 000431903805079

• Tumor Molecular Imaging with Nanoparticles ENGINEERING Cheng, Z., Yan, X., Sun, X., Shen, B., Gambhir, S. 2016; 2 (1): 132–40

  View details for DOI 10.1016/J.ENG.2016.01.027

  View details for Web of Science ID 000389355500031

• Pilot Preclinical and Clinical Evaluation of (4S)-4-(3-[18F]Fluoropropyl)-L-Glutamate (18F-FSPG) for PET/CT Imaging of Intracranial Malignancies. PloS one Mittra, E. S., Koglin, N., Mosci, C., Kumar, M., Hoehne, A., Keu, K. V., Iagaru, A. H., Mueller, A., Berndt, M., Bullich, S., Friebe, M., Schmitt-Willich, H., Gekeler, V., Fels, L. M., Bacher-Stier, C., Moon, D. H., Chin, F. T., Stephens, A. W., Dinkelborg, L. M., Gambhir, S. S. 2016; 11 (2)

  Abstract

  (S)-4-(3-[18F]Fluoropropyl)-L-glutamic acid (18F-FSPG) is a novel radiopharmaceutical for Positron Emission Tomography (PET) imaging. It is a glutamate analogue that can be used to measure xC- transporter activity. This study was performed to assess the feasibility of 18F-FSPG for imaging orthotopic brain tumors in small animals and the translation of this approach in human subjects with intracranial malignancies.For the small animal study, GS9L glioblastoma cells were implanted into brains of Fischer rats and studied with 18F-FSPG, the 18F-labeled glucose derivative 18F-FDG and with the 18F-labeled amino acid derivative 18F-FET. For the human study, five subjects with either primary or metastatic brain cancer were recruited (mean age 50.4 years). After injection of 300 MBq of 18F-FSPG, 3 whole-body PET/Computed Tomography (CT) scans were obtained and safety parameters were measured. The three subjects with brain metastases also had an 18F-FDG PET/CT scan. Quantitative and qualitative comparison of the scans was performed to assess kinetics, biodistribution, and relative efficacy of the tracers.In the small animals, the orthotopic brain tumors were visualized well with 18F-FSPG. The high tumor uptake of 18F-FSPG in the GS9L model and the absence of background signal led to good tumor visualization with high contrast (tumor/brain ratio: 32.7). 18F-FDG and 18F-FET showed T/B ratios of 1.7 and 2.8, respectively. In the human pilot study, 18F-FSPG was well tolerated and there was similar distribution in all patients. All malignant lesions were positive with 18F-FSPG except for one low-grade primary brain tumor. In the 18F-FSPG-PET-positive tumors a similar T/B ratio was observed as in the animal model.18F-FSPG is a novel PET radiopharmaceutical that demonstrates good uptake in both small animal and human studies of intracranial malignancies. Future studies on larger numbers of subjects and a wider array of brain tumors are planned.ClinicalTrials.gov NCT01186601.

  View details for DOI 10.1371/journal.pone.0148628

  View details for PubMedID 26890637

  View details for PubMedCentralID PMC4758607

• A Cystine Knot Peptide Targeting Integrin αvβ6 for Photoacoustic and Fluorescence Imaging of Tumors in Living Subjects. Journal of Nuclear Medicine Zhang, C., Kimura, R., Abou-Elkacem, L., Levi, J., Xu, L., Gambhir, S. S. 2016: 1629-1634
• AshwaMAX and Withaferin A inhibits gliomas in cellular and murine orthotopic models. Journal of neuro-oncology Chang, E., Pohling, C., Natarajan, A., Witney, T. H., Kaur, J., Xu, L., Gowrishankar, G., D'Souza, A. L., Murty, S., Schick, S., Chen, L., Wu, N., Khaw, P., Mischel, P., Abbasi, T., Usmani, S., Mallick, P., Gambhir, S. S. 2016; 126 (2): 253-64

  Abstract

  Glioblastoma multiforme (GBM) is an aggressive, malignant cancer Johnson and O'Neill (J Neurooncol 107: 359-364, 2012). An extract from the winter cherry plant (Withania somnifera ), AshwaMAX, is concentrated (4.3 %) for Withaferin A; a steroidal lactone that inhibits cancer cells Vanden Berghe et al. (Cancer Epidemiol Biomark Prev 23: 1985-1996, 2014). We hypothesized that AshwaMAX could treat GBM and that bioluminescence imaging (BLI) could track oral therapy in orthotopic murine models of glioblastoma. Human parietal-cortical glioblastoma cells (GBM2, GBM39) were isolated from primary tumors while U87-MG was obtained commercially. GBM2 was transduced with lentiviral vectors that express Green Fluorescent Protein (GFP)/firefly luciferase fusion proteins. Mutational, expression and proliferative status of GBMs were studied. Intracranial xenografts of glioblastomas were grown in the right frontal regions of female, nude mice (n = 3-5 per experiment). Tumor growth was followed through BLI. Neurosphere cultures (U87-MG, GBM2 and GBM39) were inhibited by AshwaMAX at IC50 of 1.4, 0.19 and 0.22 µM equivalent respectively and by Withaferin A with IC50 of 0.31, 0.28 and 0.25 µM respectively. Oral gavage, every other day, of AshwaMAX (40 mg/kg per day) significantly reduced bioluminescence signal (n = 3 mice, p < 0.02, four parameter non-linear regression analysis) in preclinical models. After 30 days of treatment, bioluminescent signal increased suggesting onset of resistance. BLI signal for control, vehicle-treated mice increased and then plateaued. Bioluminescent imaging revealed diffuse growth of GBM2 xenografts. With AshwaMAX, GBM neurospheres collapsed at nanomolar concentrations. Oral treatment studies on murine models confirmed that AshwaMAX is effective against orthotopic GBM. AshwaMAX is thus a promising candidate for future clinical translation in patients with GBM.

  View details for DOI 10.1007/s11060-015-1972-1

  View details for PubMedID 26650066

• Engineered PD-1 variants as immunotherapies for cancer Gordon, S. R., Maute, R., Mayer, A., McCracken, M., Natarajan, A., Guo, N., Kimura, R., Tsai, J. M., Manglik, A., Kruse, A., Gambhir, S., Weissman, I. L., Ring, A. M. AMER ASSOC CANCER RESEARCH. 2016

  View details for DOI 10.1158/2326-6074.CRICIMTEATIAACR15-B101

  View details for Web of Science ID 000375484400088

• Artificial MicroRNAs as Novel Secreted Reporters for Cell Monitoring in Living Subjects. PloS one Ronald, J. A., D'Souza, A. L., Chuang, H., Gambhir, S. S. 2016; 11 (7)

  Abstract

  Reporter genes are powerful technologies that can be used to directly inform on the fate of transplanted cells in living subjects. Imaging reporter genes are often employed to quantify cell number, location(s), and viability with various imaging modalities. To complement this, reporters that are secreted from cells can provide a low-cost, in vitro diagnostic test to monitor overall cell viability at relatively high frequency without knowing the locations of all cells. Whereas protein-based secretable reporters have been developed, an RNA-based reporter detectable with amplification inherent PCR-based assays has not been previously described. MicroRNAs (miRNAs) are short non-coding RNAs (18-22 nt) that regulate mRNA translation and are being explored as relatively stable blood-based disease biomarkers. We developed an artificial miRNA-based secreted reporter, called Sec-miR, utilizing a coding sequence that is not expressed endogenously and does not have any known vertebrate target. Sec-miR was detectable in both the cells and culture media of transiently transfected cells. Cells stably expressing Sec-miR also reliably secreted it into the culture media. Mice implanted with parental HeLa cells or HeLa cells expressing both Sec-miR and the bioluminescence imaging (BLI) reporter gene Firefly luciferase (FLuc) were monitored over time for tumor volume, FLuc signal via BLI, and blood levels of Sec-miR. Significantly (p<0.05) higher Sec-miR was found in the blood of mice bearing Sec-miR-expressing tumors compared to parental cell tumors at 21 and 28 days after implantation. Importantly, blood Sec-miR reporter levels after day 21 showed a trend towards correlation with tumor volume (R2 = 0.6090; p = 0.0671) and significantly correlated with FLuc signal (R2 = 0.7067; p<0.05). Finally, we could significantly (p<0.01) amplify Sec-miR secretion into the cell media by chaining together multiple Sec-miR copies (4 instead of 1 or 2) within an expression cassette. Overall, we show that a novel complement of BLI together with a unique Sec-miR reporter adds an in vitro RNA-based diagnostic to enhance the monitoring of transplanted cells. While Sec-miR was not as sensitive as BLI for monitoring cell number, it may be more sensitive than clinically-relevant positron emission tomography (PET) reporter assays. Future work will focus on improving cell detectability via improved secretion of Sec-miR reporters from cells and more sensitive detection platforms, as well as, exploring other miRNA sequences to allow multiplexed monitoring of more than one cell population at a time. Continued development may lead to more refined and precise monitoring of cell-based therapies.

  View details for DOI 10.1371/journal.pone.0159369

  View details for PubMedID 27442530

  View details for PubMedCentralID PMC4956193

• Comparison of Deconvolution Filters for Photoacoustic Tomography. PloS one Van de Sompel, D., Sasportas, L. S., Jokerst, J. V., Gambhir, S. S. 2016; 11 (3)

  Abstract

  In this work, we compare the merits of three temporal data deconvolution methods for use in the filtered backprojection algorithm for photoacoustic tomography (PAT). We evaluate the standard Fourier division technique, the Wiener deconvolution filter, and a Tikhonov L-2 norm regularized matrix inversion method. Our experiments were carried out on subjects of various appearances, namely a pencil lead, two man-made phantoms, an in vivo subcutaneous mouse tumor model, and a perfused and excised mouse brain. All subjects were scanned using an imaging system with a rotatable hemispherical bowl, into which 128 ultrasound transducer elements were embedded in a spiral pattern. We characterized the frequency response of each deconvolution method, compared the final image quality achieved by each deconvolution technique, and evaluated each method's robustness to noise. The frequency response was quantified by measuring the accuracy with which each filter recovered the ideal flat frequency spectrum of an experimentally measured impulse response. Image quality under the various scenarios was quantified by computing noise versus resolution curves for a point source phantom, as well as the full width at half maximum (FWHM) and contrast-to-noise ratio (CNR) of selected image features such as dots and linear structures in additional imaging subjects. It was found that the Tikhonov filter yielded the most accurate balance of lower and higher frequency content (as measured by comparing the spectra of deconvolved impulse response signals to the ideal flat frequency spectrum), achieved a competitive image resolution and contrast-to-noise ratio, and yielded the greatest robustness to noise. While the Wiener filter achieved a similar image resolution, it tended to underrepresent the lower frequency content of the deconvolved signals, and hence of the reconstructed images after backprojection. In addition, its robustness to noise was poorer than that of the Tikhonov filter. The performance of the Fourier filter was found to be the poorest of all three methods, based on the reconstructed images' lowest resolution (blurriest appearance), generally lowest contrast-to-noise ratio, and lowest robustness to noise. Overall, the Tikhonov filter was deemed to produce the most desirable image reconstructions.

  View details for DOI 10.1371/journal.pone.0152597

  View details for PubMedID 27031832

  View details for PubMedCentralID PMC4816281

• AshwaMAX and Withaferin A inhibits gliomas in cellular and murine orthotopic models JOURNAL OF NEURO-ONCOLOGY Chang, E., Pohling, C., Natarajan, A., Witney, T. H., Kaur, J., Xu, L., Gowrishankar, G., D'Souza, A. L., Murty, S., Schick, S., Chen, L., Wu, N., Khaw, P., Mischel, P., Abbasi, T., Usmani, S., Mallick, P., Gambhir, S. S. 2016; 126 (2): 253-264

  Abstract

  Glioblastoma multiforme (GBM) is an aggressive, malignant cancer Johnson and O'Neill (J Neurooncol 107: 359-364, 2012). An extract from the winter cherry plant (Withania somnifera ), AshwaMAX, is concentrated (4.3 %) for Withaferin A; a steroidal lactone that inhibits cancer cells Vanden Berghe et al. (Cancer Epidemiol Biomark Prev 23: 1985-1996, 2014). We hypothesized that AshwaMAX could treat GBM and that bioluminescence imaging (BLI) could track oral therapy in orthotopic murine models of glioblastoma. Human parietal-cortical glioblastoma cells (GBM2, GBM39) were isolated from primary tumors while U87-MG was obtained commercially. GBM2 was transduced with lentiviral vectors that express Green Fluorescent Protein (GFP)/firefly luciferase fusion proteins. Mutational, expression and proliferative status of GBMs were studied. Intracranial xenografts of glioblastomas were grown in the right frontal regions of female, nude mice (n = 3-5 per experiment). Tumor growth was followed through BLI. Neurosphere cultures (U87-MG, GBM2 and GBM39) were inhibited by AshwaMAX at IC50 of 1.4, 0.19 and 0.22 µM equivalent respectively and by Withaferin A with IC50 of 0.31, 0.28 and 0.25 µM respectively. Oral gavage, every other day, of AshwaMAX (40 mg/kg per day) significantly reduced bioluminescence signal (n = 3 mice, p < 0.02, four parameter non-linear regression analysis) in preclinical models. After 30 days of treatment, bioluminescent signal increased suggesting onset of resistance. BLI signal for control, vehicle-treated mice increased and then plateaued. Bioluminescent imaging revealed diffuse growth of GBM2 xenografts. With AshwaMAX, GBM neurospheres collapsed at nanomolar concentrations. Oral treatment studies on murine models confirmed that AshwaMAX is effective against orthotopic GBM. AshwaMAX is thus a promising candidate for future clinical translation in patients with GBM.

  View details for DOI 10.1007/s11060-015-1972-1

  View details for Web of Science ID 000368728300005

• Multiscale Framework for Imaging Radiolabeled Therapeutics. Molecular pharmaceutics Natarajan, A., Türkcan, S., Gambhir, S. S., Pratx, G. 2015; 12 (12): 4554-4560

  Abstract

  The resistance of a tumor to a drug is the result of bulk properties of the tumor tissue as well as phenotypic variations displayed by single cells. Here, we show that radioisotopic detection methods, commonly used for tracking the tissue distribution of drug compounds, can be extended to the single-cell level to image the same molecule over a range of physical scales. The anticancer drug rituximab was labeled with short-lived radionuclides ((89)Zr/(64)Cu) and its accumulation at the organ level was imaged using PET in a humanized transgenic mouse model of non-Hodgkin's lymphoma. To capture the distribution of the drug at a finer scale, tissue sections and single living cells were imaged using radioluminescence microscopy (RLM), a novel method that can detect radionuclides with single-cell resolution. In vivo PET images (24 h postinjection) showed that [(89)Zr]rituximab targeted the intended site of human CD20 expression, the spleen. Within this organ, RLM was used to resolve radiotracer accumulation in the splenic red pulp. In a separate study, RLM highlighted marked differences between single cells, with binding of the radiolabeled antibody ranging from background levels to 1200 radionuclides per cell. Overall, RLM images demonstrated significantly higher spatial resolution and sensitivity than conventional storage-phosphor autoradiography. In conclusion, this combination of PET and RLM provides a unique opportunity for exploring the molecular mechanism of drugs by tracking the same molecule over multiple physical scales, ranging from single living cells to organs substructures and entire living subjects.

  View details for DOI 10.1021/acs.molpharmaceut.5b00392

  View details for PubMedID 26460685

• Dynamic Noninvasive Genomic Monitoring for Outcome Prediction in Diffuse Large B-Cell Lymphoma Kurtz, D. M., Scherer, F., Newman, A. M., Lovejoy, A. F., Klass, D. M., Chabon, J. J., Gambhir, S., Diehn, M., Alizadeh, A. A. AMER SOC HEMATOLOGY. 2015

  View details for Web of Science ID 000368019000194

• Prospective Comparison of 99mTc-MDP Scintigraphy, Combined 18F-NaF and 18F-FDG PET/CT, and Whole-Body MRI in Patients with Breast and Prostate Cancer. Journal of nuclear medicine Minamimoto, R., Loening, A., Jamali, M., Barkhodari, A., Mosci, C., Jackson, T., Obara, P., Taviani, V., Gambhir, S. S., Vasanawala, S., Iagaru, A. 2015; 56 (12): 1862-1868

  Abstract

  We prospectively evaluated the combined (18)F-NaF/(18)F-FDG PET/CT in patients with breast and prostate cancers, and compared the results to (99m)Tc MDP bone scintigraphy (BS) and whole-body MRI (WBMRI).30 patients (15 women with breast cancer and 15 men with prostate cancer) referred for standard of care BS were prospectively enrolled in this study. (18)F-NaF/(18)F-FDG PET/CT and WBMRI were performed following BS. WBMRI protocol consisted of both non-contrast enhanced and contrast enhanced sequences. Lesions detected with each test were tabulated and the results were compared.For extra skeletal lesions, (18)F-/(18)F-FDG PET/CT and WBMRI had no statistically significant differences in sensitivity (92.9% vs 92.9%, P = 1.00), PPV (81.3% vs 86.7%, P = 0.68) and accuracy (76.5% vs 82.4%, P = 0.56). However, (18)F-/(18)F-FDG PET/CT showed significantly higher sensitivity and accuracy than WBMRI (96.2% vs 81.4%, P<0.001, 89.8% vs 74.7%, P = 0.01) and BS (96.2% vs 64.6%, P<0.001, 89.8% vs 65.9%, P<0.001) for the detection of skeletal lesions. Overall, (18)F-/(18)F-FDG PET/CT showed higher sensitivity and accuracy than WBMRI (95.7% vs 83.3%, P<0.002, 87.6% vs 76.0%, P< 0.02), but not statistically significant when compared to a combination of WBMRI and BS (95.7% vs 91.6%, P = 0.17, 87.6% vs 83.0%, P = 0.53). (18)F-/(18)F-FDG PET/CT showed no significant difference with a combination of (18)F-/(18)F-FDG PET/CT and WBMRI. No statistically significant differences in PPV were noted among the 3 examinations.The (18)F NaF/(18)F FDG PET/CT is superior to WBMRI and (99m)Tc-MDP scintigraphy for evaluation of skeletal disease extent. Further, (18)F NaF/(18)F FDG PET/CT and WBMRI detected extra-skeletal disease that may change the management of these patients. The (18)F NaF/(18)F FDG PET/CT provide similar diagnostic ability with combination of WBMRI and BS in patients with breast and prostate cancers. Larger cohorts are needed in order to confirm these preliminary findings, ideally using the newly introduced simultaneous PET/MRI scanners.

  View details for DOI 10.2967/jnumed.115.162610

  View details for PubMedID 26405167

• Multiscale Framework for Imaging Radio labeled Therapeutics MOLECULAR PHARMACEUTICS Natarajan, A., Tuerkcan, S., Gambhir, S. S., Pratx, G. 2015; 12 (12): 4554-4560

  View details for DOI 10.1021/acs.molpharmaceur.5b00392

  View details for Web of Science ID 000366151500036

  View details for PubMedID 26460685

• Further validation to support clinical translation of [(18)F]FTC-146 for imaging sigma-1 receptors. EJNMMI research Shen, B., James, M. L., Andrews, L., Lau, C., Chen, S., Palner, M., Miao, Z., Arksey, N. C., Shuhendler, A. J., Scatliffe, S., Kaneshige, K., Parsons, S. M., McCurdy, C. R., Salehi, A., Gambhir, S. S., Chin, F. T. 2015; 5 (1): 49-?

  Abstract

  This study aims to further evaluate the specificity and selectivity of [(18)F]FTC-146 and obtain additional data to support its clinical translation.The binding of [(19)F]FTC-146 to vesicular acetylcholine transporter (VAChT) was evaluated using [(3)H]vesamicol and PC12(A123.7) cells in an in vitro binding assay. The uptake and kinetics of [(18)F]FTC-146 in S1R-knockout mice (S1R-KO) compared to wild-type (WT) littermates was assessed using dynamic positron emission tomography (PET) imaging. Ex vivo autoradiography and histology were conducted using a separate cohort of S1R-KO/WT mice, and radiation dosimetry was calculated from WT mouse data (extrapolated for human dosing). Toxicity studies in Sprague-Dawley rats were performed with a dose equivalent to 250× the anticipated clinical dose of [(19)F]FTC-146 mass.VAChT binding assay results verified that [(19)F]FTC-146 displays negligible affinity for VAChT (K i = 450 ± 80 nM) compared to S1R. PET images demonstrated significantly higher tracer uptake in WT vs. S1R-KO brain (4.57 ± 1.07 vs. 1.34 ± 0.4 %ID/g at 20-25 min, n = 4, p < 0.05). In S1R-KO mice, it was shown that rapid brain uptake and clearance 10 min post-injection, which are consistent with previous S1R-blocking studies in mice. Three- to fourfold higher tracer uptake was observed in WT relative to S1R-KO mouse brains by ex vivo autoradiography. S1R staining coincided well with the autoradiographic data in all examined brain regions (r (2) = 0.85-0.95). Biodistribution results further demonstrated high [(18)F]FTC-146 accumulation in WT relative to KO mouse brain and provided quantitative information concerning tracer uptake in S1R-rich organs (e.g., heart, lung, pancreas) for WT mice vs. age-matched S1R-KO mice. The maximum allowed dose per scan in humans as extrapolated from mouse dosimetry was 33.19 mCi (1228.03 MBq). No significant toxicity was observed even at a 250X dose of the maximum carrier mass [(19)F]FTC-146 expected to be injected for human studies.Together, these data indicate that [(18)F]FTC-146 binds specifically to S1Rs and is a highly promising radiotracer ready for clinical translation to investigate S1R-related diseases.

  View details for DOI 10.1186/s13550-015-0122-2

  View details for PubMedID 26384292

  View details for PubMedCentralID PMC4573970

• Photoacoustic Tomography Detects Early Vessel Regression and Normalization During Ovarian Tumor Response to the Antiangiogenic Therapy Trebananib. Journal of nuclear medicine Bohndiek, S. E., Sasportas, L. S., Machtaler, S., Jokerst, J. V., Hori, S., Gambhir, S. S. 2015; 56 (12): 1942-1947

  Abstract

  The primary aim of this study was to assess the potential of in vivo photoacoustic tomography (PAT) for direct functional measurement of ovarian tumor response to anti-angiogenic therapy.In vivo studies were performed with institutional animal care and use committee approval. We used an orthotopic mouse model of ovarian cancer treated with Trebananib (n = 9) or vehicle (n = 9). Tumor-bearing mice were randomized into Trebananib or vehicle groups at day 10 and dosed on days 12, 15 and 18 post implantation. PAT and blood draws were performed at day 10, then 24 hours after each drug dose. Tumors were excised for histopathology following the final studies on day 19. Data analysis to test for statistical significance was performed blinded.Blockade of angiopoietin signaling using Trebananib resulted in reduced total hemoglobin-weighted PA signal (n = 9, P = 0.01) and increased oxyhemoglobin-weighted PA signal (n = 9, P<0.01). The latter observation indicated normalization of the residual tumor vessels, which was also implied by low levels of angiopoietin 1 in serum biomarker profiling (0.76±0.12ng/mL). These non-invasive measures reflected a 30% reduction in microvessel density and increased vessel maturation in ex vivo sections.PAT is able to evaluate both vessel regression and normalization in response to Trebananib. Non-invasive imaging data was supported by modulation of serum markers in vitro and ex vivo histopathology.

  View details for DOI 10.2967/jnumed.115.160002

  View details for PubMedID 26315834

• (18)F-FPRGD2 PET/CT imaging of musculoskeletal disorders. Annals of nuclear medicine Withofs, N., Charlier, E., Simoni, P., Alvarez-Miezentseva, V., Mievis, F., Giacomelli, F., Mella, C., Gambhir, S. S., Malaise, O., de Seny, D., Malaise, M., Hustinx, R. 2015; 29 (10): 839-847

  Abstract

  This work reports on musculoskeletal uptake of (18)F-FPRGD2, targeting the integrin αvβ3, in patients who had undergone (18)F-FPRGD2 positron emission tomography combined with computed tomography (PET/CT) for oncologic purposes.Whole-body (18)F-FPRGD2 PET/CT images of 62 cancer patients were retrospectively reviewed to detect foci of musculoskeletal (18)F-FPRGD2 uptake. For 37 patients, a FDG PET/CT performed in clinical settings was available. In each joint with an abnormal uptake, the maximum standardized uptake value (SUVmax) was estimated.A total of 260 musculoskeletal foci of (18)F-FPRGD2 uptake were detected. Most common sites of uptake were joints and discs (n = 160; 61.5 %), entheses (osteotendinous and osteoligamentous junctions; n = 55; 21.2 %) and recent fractures (n = 18; 6.9 %). In addition, 27 (10.4 %) miscellaneous foci were detected. Out of the 146 lesions for which a FDG PET was available, 63 % showed both (18)F-FPRGD2 and FDG uptake, 33.6 % did not show FDG avidity and 3.4 % showed only FDG uptake. The uptake intensity of the 92 lesions positive with (18)F-FPRGD2 and FDG was similar with both radiopharmaceuticals, but the target-to-background (blood pool or muscle) ratios were significantly higher with (18)F-FPRGD2 than with FDG (p < 0.0001).The (18)F-FPRGD2 uptake in joints, spine degenerative diseases and tendons was highly prevalent in our population. Up to one-third of (18)F-FPRGD2 foci showed no FDG uptake suggesting that (18)F-FPRGD2 signal may not be related to inflammatory angiogenesis only.

  View details for DOI 10.1007/s12149-015-1011-5

  View details for PubMedID 26254227

• Imaging patients with breast and prostate cancers using combined 18F NaF/18F FDG and TOF simultaneous PET/ MRI. EJNMMI physics Iagaru, A., Minamimoto, R., Jamali, M., Barkodhodari, A., Gambhir, S. S., Vasanawala, S. 2015; 2: A65-?

  View details for DOI 10.1186/2197-7364-2-S1-A65

  View details for PubMedID 26956325

  View details for PubMedCentralID PMC4798635

• Engineering high-affinity PD-1 variants for optimized immunotherapy and immuno-PET imaging. Proceedings of the National Academy of Sciences of the United States of America Maute, R. L., Gordon, S. R., Mayer, A. T., McCracken, M. N., Natarajan, A., Ring, N. G., Kimura, R., Tsai, J. M., Manglik, A., Kruse, A. C., Gambhir, S. S., Weissman, I. L., Ring, A. M. 2015; 112 (47): E6506-14

  Abstract

  Signaling through the immune checkpoint programmed cell death protein-1 (PD-1) enables tumor progression by dampening antitumor immune responses. Therapeutic blockade of the signaling axis between PD-1 and its ligand programmed cell death ligand-1 (PD-L1) with monoclonal antibodies has shown remarkable clinical success in the treatment of cancer. However, antibodies have inherent limitations that can curtail their efficacy in this setting, including poor tissue/tumor penetrance and detrimental Fc-effector functions that deplete immune cells. To determine if PD-1:PD-L1-directed immunotherapy could be improved with smaller, nonantibody therapeutics, we used directed evolution by yeast-surface display to engineer the PD-1 ectodomain as a high-affinity (110 pM) competitive antagonist of PD-L1. In contrast to anti-PD-L1 monoclonal antibodies, high-affinity PD-1 demonstrated superior tumor penetration without inducing depletion of peripheral effector T cells. Consistent with these advantages, in syngeneic CT26 tumor models, high-affinity PD-1 was effective in treating both small (50 mm(3)) and large tumors (150 mm(3)), whereas the activity of anti-PD-L1 antibodies was completely abrogated against large tumors. Furthermore, we found that high-affinity PD-1 could be radiolabeled and applied as a PET imaging tracer to efficiently distinguish between PD-L1-positive and PD-L1-negative tumors in living mice, providing an alternative to invasive biopsy and histological analysis. These results thus highlight the favorable pharmacology of small, nonantibody therapeutics for enhanced cancer immunotherapy and immune diagnostics.

  View details for DOI 10.1073/pnas.1519623112

  View details for PubMedID 26604307

  View details for PubMedCentralID PMC4664306

• Glioblastoma Multiforme Recurrence: An Exploratory Study of F-18 FPPRGD(2) PET/CT1 RADIOLOGY Iagaru, A., Mosci, C., Mittra, E., Zaharchuk, G., Fischbein, N., Harsh, G., Li, G., Nagpal, S., Recht, L., Gambhir, S. S. 2015; 277 (2): 497-506

  Abstract

  Purpose To prospectively evaluate fluorine 18 ((18)F) 2-fluoropropionyl-labeled PEGylated dimeric arginine-glycine-aspartic acid (RGD) peptide (PEG3-E[c{RGDyk}]2) (FPPRGD2) positron emission tomography (PET) in patients with glioblastoma multiforme (GBM). Materials and Methods The institutional review board approved this HIPAA-compliant protocol. Written informed consent was obtained from each patient. (18)F FPPRGD2 uptake was measured semiquantitatively in the form of maximum standardized uptake values (SUVmax) and uptake volumes before and after treatment with bevacizumab. Vital signs and laboratory results were collected before, during, and after the examinations. A nonparametric version of multivariate analysis of variance was used to assess safety outcome measures simultaneously across time points. A paired two-sample t test was performed to compare SUVmax. Results A total of 17 participants (eight men, nine women; age range, 25-65 years) were enrolled prospectively. (18)F FPPRGD2 PET/computed tomography (CT), (18)F fluorodeoxyglucose (FDG) PET/CT, and brain magnetic resonance (MR) imaging were performed within 3 weeks, prior to the start of bevacizumab therapy. In eight of the 17 patients (47%), (18)F FPPRGD2 PET/CT was repeated 1 week after the start of bevacizumab therapy; six patients (35%) underwent (18)F FPPRGD2 PET/CT a third time 6 weeks after starting bevacizumab therapy. There were no changes in vital signs, electrocardiographic findings, or laboratory values that qualified as adverse events. One patient (6%) had recurrent GBM identified only on (18)F FPPRGD2 PET images, and subsequent MR images enabled confirmation of recurrence. Of the 17 patients, 14 (82%) had recurrent GBM identified on (18)F FPPRGD2 PET and brain MR images, while (18)F FDG PET enabled identification of recurrence in 13 (76%) patients. Two patients (12%) had no recurrent GBM. Conclusion (18)F FPPRGD2 is a safe PET radiopharmaceutical that has increased uptake in GBM lesions. Larger cohorts are required to confirm these preliminary findings. (©) RSNA, 2015 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.2015141550

  View details for Web of Science ID 000368435100026

• Biodistribution of the (18)F-FPPRGD2 PET radiopharmaceutical in cancer patients: an atlas of SUV measurements. European journal of nuclear medicine and molecular imaging Minamimoto, R., Jamali, M., Barkhodari, A., Mosci, C., Mittra, E., Shen, B., Chin, F., Gambhir, S. S., Iagaru, A. 2015; 42 (12): 1850-1858

  Abstract

  The aim of this study was to investigate the biodistribution of 2-fluoropropionyl-labeled PEGylated dimeric arginine-glycine-aspartic acid (RGD) peptide (PEG3-E[c{RGDyk}]2) ((18)F-FPPRGD2) in cancer patients and to compare its uptake in malignant lesions with (18)F-FDG uptake.A total of 35 patients (11 men, 24 women, mean age 52.1 ± 10.8 years) were enrolled prospectively and had (18)F-FPPRGD2 PET/CT prior to treatment. Maximum standardized uptake values (SUVmax) and mean SUV (SUVmean) were measured in 23 normal tissues in each patient, as well as in known or suspected cancer lesions. Differences between (18)F-FPPRGD2 uptake and (18)F-FDG uptake were also evaluated in 28 of the 35 patients.Areas of high (18)F-FPPRGD2 accumulation (SUVmax range 8.9 - 94.4, SUVmean range 7.1 - 64.4) included the bladder and kidneys. Moderate uptake (SUVmax range 2.1 - 6.3, SUVmean range 1.1 - 4.5) was found in the choroid plexus, salivary glands, thyroid, liver, spleen, pancreas, small bowel and skeleton. Compared with (18)F-FDG, (18)F-FPPRGD2 showed higher tumor-to-background ratio in brain lesions (13.4 ± 8.5 vs. 1.1 ± 0.5, P < 0.001), but no significant difference in body lesions (3.2 ± 1.9 vs. 4.4 ± 4.2, P = 0.10). There was no significant correlation between the uptake values (SUVmax and SUVmean) for (18)F FPPRGD2 and those for (18)F-FDG.The biodistribution of (18)F-FPPRGD2 in cancer patients is similar to that of other RGD dimer peptides and it is suitable for clinical use. The lack of significant correlation between (18)F-FPPRGD2 and (18)F-FDG uptake confirms that the information provided by each PET tracer is different.

  View details for DOI 10.1007/s00259-015-3096-4

  View details for PubMedID 26062933

• Novel Radiotracer for ImmunoPET Imaging of PD-1 Checkpoint Expression on Tumor Infiltrating Lymphocytes. Bioconjugate chemistry Natarajan, A., Mayer, A. T., Xu, L., Reeves, R. E., Gano, J., Gambhir, S. S. 2015; 26 (10): 2062-2069

  Abstract

  Immune checkpoint signaling through the programmed death 1 (PD-1) axis to its ligand (PD-L1) significantly dampens anti-tumor immune responses. Cancer patients treated with checkpoint inhibitors that block this suppressive signaling have exhibited objective response rates of 20-40% for advanced solid tumors, lymphomas, and malignant melanomas. This represents a tremendous advance in cancer treatment. Unfortunately, all patients do not respond to immune checkpoint blockade. Recent findings suggest that patients with tumor infiltrating lymphocytes (TILs) expressing PD-1 may be most likely to respond to αPD-1/PD-L1 checkpoint inhibitors. There is a compelling need for diagnostic and prognostic imaging tools to assess the PD-1 status of TILs in vivo. Here we have developed a novel immunoPET tracer to image PD-1 expressing TILs in a transgenic mouse model bearing melanoma. A (64)Cu labeled anti-mouse antibody (IgG) PD-1 immuno positron emission tomography (PET) tracer was developed to detect PD-1 expressing murine TILs. Quality control of the tracer showed >95% purity by HPLC and >70% immunoreactivity in an in vitro cell binding assay. ImmunoPET scans were performed over 1-48 h on Foxp3+.LuciDTR4 mice bearing B16-F10 melanoma tumors. Mice receiving anti-PD-1 tracer (200 ± 10 μCi/10-12 μg/200 μL) revealed high tracer uptake in lymphoid organs and tumors. BLI images of FoxP3(+) CD4(+) Tregs known to express PD-1 confirmed lymphocyte infiltration of tumors at the time of PET imaging. Biodistribution measurements performed at 48 h revealed a high (11×) tumor to muscle uptake ratio of the PET tracer (p < 0.05). PD-1 tumors exhibited 7.4 ± 0.7%ID/g tracer uptake and showed a 2× fold signal decrease when binding was blocked by unlabeled antibody. To the best of our knowledge this data is the first report to image PD-1 expression in living subjects with PET. This radiotracer has the potential to assess the prognostic value of PD-1 in preclinical models of immunotherapy and may ultimately aid in predicting response to therapies targeting immune checkpoints.

  View details for DOI 10.1021/acs.bioconjchem.5b00318

  View details for PubMedID 26307602

• PET imaging of tumor glycolysis downstream of hexokinase through noninvasive measurement of pyruvate kinase M2. Science translational medicine Witney, T. H., James, M. L., Shen, B., Chang, E., Pohling, C., Arksey, N., Hoehne, A., Shuhendler, A., Park, J., Bodapati, D., Weber, J., Gowrishankar, G., Rao, J., Chin, F. T., Gambhir, S. S. 2015; 7 (310): 310ra169-?

  Abstract

  Cancer cells reprogram their metabolism to meet increased biosynthetic demands, commensurate with elevated rates of replication. Pyruvate kinase M2 (PKM2) catalyzes the final and rate-limiting step in tumor glycolysis, controlling the balance between energy production and the synthesis of metabolic precursors. We report here the synthesis and evaluation of a positron emission tomography (PET) radiotracer, [(11)C]DASA-23, that provides a direct noninvasive measure of PKM2 expression in preclinical models of glioblastoma multiforme (GBM). In vivo, orthotopic U87 and GBM39 patient-derived tumors were clearly delineated from the surrounding normal brain tissue by PET imaging, corresponding to exclusive tumor-associated PKM2 expression. In addition, systemic treatment of mice with the PKM2 activator TEPP-46 resulted in complete abrogation of the PET signal in intracranial GBM39 tumors. Together, these data provide the basis for the clinical evaluation of imaging agents that target this important gatekeeper of tumor glycolysis.

  View details for DOI 10.1126/scitranslmed.aac6117

  View details for PubMedID 26491079

• Combined F-18-NaF and F-18-FDG PET/CT in the Evaluation of Sarcoma Patients CLINICAL NUCLEAR MEDICINE Jackson, T., Mosci, C., von Eyben, R., Mittra, E., Ganjoo, K., Biswal, S., Gambhir, S. S., Iagaru, A. 2015; 40 (9): 720-724

  Abstract

  The combined administration of F-NaF and F-FDG in a single PET/CT scan has the potential to improve patient convenience and cancer detection. Here we report the use of this approach for patients with sarcomas.This is a retrospective review of 21 patients (12 men, 9 women; age, 19-66 years) with biopsy-proven sarcomas who had separate F-NaF PET/CT, F-FDG PET/CT, and combined F-NaF/F-FDG PET/CT scans for evaluation of malignancy. Two board-certified nuclear medicine physicians and 1 board-certified musculoskeletal radiologist were randomly assigned to review the scans. Results were analyzed for sensitivity and specificity, using linear regression and receiver operating characteristics.A total of 13 patients had metastatic disease on F-NaF PET/CT, F-FDG PET/CT, and combined F-NaF/F-FDG PET/CT. Skeletal disease was more extensive on the F-NaF PET/CT scan than on the F-FDG PET/CT in 3 patients, whereas in 1 patient, F-FDG PET/CT showed skeletal disease and the F-NaF PET/CT was negative. Extraskeletal lesions were detected on both F-FDG and combined F-NaF/F-FDG PET/CT in 20 patients, with 1 discordant finding in the lung.The combined F-NaF/F-FDG PET/CT scan allows for accurate evaluation of sarcoma patients. Further evaluation of this proposed imaging modality is warranted to identify the most suitable clinical scenarios, including initial treatment strategy and evaluation of response to therapy.

  View details for DOI 10.1097/RLU.0000000000000845

  View details for Web of Science ID 000359668600005

  View details for PubMedID 26053718

• 18F-FDG PET Imaging Utilization in the National Lung Screening Trial Nair, V. S., Sundaram, V., Gould, M. K., Gambhir, S. S., Desai, M. ELSEVIER SCIENCE INC. 2015: S396–S397

  View details for Web of Science ID 000370365101407

• A Systematic Comparison of 18F-C-SNAT to Established Radiotracer Imaging Agents for the Detection of Tumor Response to Treatment. Clinical cancer research Witney, T. H., Hoehne, A., Reeves, R. E., Ilovich, O., Namavari, M., Shen, B., Chin, F. T., Rao, J., Gambhir, S. S. 2015; 21 (17): 3896-3905

  Abstract

  An early readout of tumor response to therapy through measurement of drug or radiation-induced cell death may provide important prognostic indications and improved patient management. It has been shown that the uptake of (18)F-C-SNAT can be used to detect early response to therapy in tumors by positron emission tomography (PET) via a mechanism of caspase-3-triggered nanoaggregation.Here, we compared the preclinical utility of (18)F-C-SNAT for the detection of drug-induced cell death to clinically evaluated radiotracers, (18)F-FDG, (99m)Tc-Annexin V, and (18)F-ML-10 in tumor cells in culture, and in tumor-bearing mice in vivo.In drug-treated lymphoma cells, (18)F-FDG, (99m)Tc-Annexin V, and (18)F-C-SNAT cell-associated radioactivity correlated well to levels of cell death (R(2) > 0.8; P < 0.001), with no correlation measured for (18)F-ML-10 (R(2) = 0.05; P > 0.05). A similar pattern of response was observed in two human NSCLC cell lines following carboplatin treatment. EL-4 tumor uptake of (99m)Tc-Annexin V and (18)F-C-SNAT were increased 1.4- and 2.1-fold, respectively, in drug-treated versus naïve control animals (P < 0.05), although (99m)Tc-Annexin V binding did not correlate to ex vivo TUNEL staining of tissue sections. A differential response was not observed with either (18)F-FDG or (18)F-ML-10.We have demonstrated here that (18)F-C-SNAT can sensitively detect drug-induced cell death in murine lymphoma and human NSCLC. Despite favorable image contrast obtained with (18)F-C-SNAT, the development of next-generation derivatives, using the same novel and promising uptake mechanism, but displaying improved biodistribution profiles, are warranted for maximum clinical utility. Clin Cancer Res; 21(17); 3896-905. ©2015 AACR.

  View details for DOI 10.1158/1078-0432.CCR-14-3176

  View details for PubMedID 25972517

  View details for PubMedCentralID PMC4558304

• Androgen Receptor Splice Variants Dimerize to Transactivate Target Genes CANCER RESEARCH Xu, D., Zhan, Y., Qi, Y., Cao, B., Bai, S., Xu, W., Gambhir, S. S., Lee, P., Sartor, O., Flemington, E. K., Zhang, H., Hu, C., Dong, Y. 2015; 75 (17): 3663-3671

  Abstract

  Constitutively active androgen receptor splice variants (AR-V) lacking the ligand-binding domain have been implicated in the pathogenesis of castration-resistant prostate cancer and in mediating resistance to newer drugs that target the androgen axis. AR-V regulates expression of both canonical AR targets and a unique set of cancer-specific targets that are enriched for cell-cycle functions. However, little is known about how AR-V controls gene expression. Here, we report that two major AR-Vs, termed AR-V7 and AR(v567es), not only homodimerize and heterodimerize with each other but also heterodimerize with full-length androgen receptor (AR-FL) in an androgen-independent manner. We found that heterodimerization of AR-V and AR-FL was mediated by N- and C-terminal interactions and by the DNA-binding domain of each molecule, whereas AR-V homodimerization was mediated only by DNA-binding domain interactions. Notably, AR-V dimerization was required to transactivate target genes and to confer castration-resistant cell growth. Our results clarify the mechanism by which AR-Vs mediate gene regulation and provide a pivotal pathway for rational drug design to disrupt AR-V signaling as a rational strategy for the effective treatment of advanced prostate cancer.

  View details for DOI 10.1158/0008-5472.CAN-15-0381

  View details for Web of Science ID 000361917100021

  View details for PubMedID 26060018

  View details for PubMedCentralID PMC4558376

• Diketopyrrolopyrrole-Based Semiconducting Polymer Nanoparticles for In Vivo Photoacoustic Imaging. Advanced materials Pu, K., Mei, J., Jokerst, J. V., Hong, G., Antaris, A. L., Chattopadhyay, N., Shuhendler, A. J., Kurosawa, T., Zhou, Y., Gambhir, S. S., Bao, Z., Rao, J. 2015; 27 (35): 5184-5190

  Abstract

  Diketopyrrolopyrrole-based semiconducting polymer nanoparticles with high photostability and strong photoacoustic brightness are designed and synthesized, which results in 5.3-fold photoacoustic signal enhancement in tumor xenografts after systemic administration.

  View details for DOI 10.1002/adma.201502285

  View details for PubMedID 26247171

  View details for PubMedCentralID PMC4567488

• Multitarget, quantitative nanoplasmonic electrical field-enhanced resonating device (NE2RD) for diagnostics. Proceedings of the National Academy of Sciences of the United States of America Inci, F., Filippini, C., Baday, M., Ozen, M. O., Calamak, S., Durmus, N. G., Wang, S., Hanhauser, E., Hobbs, K. S., Juillard, F., Kuang, P. P., Vetter, M. L., Carocci, M., Yamamoto, H. S., Takagi, Y., Yildiz, U. H., Akin, D., Wesemann, D. R., Singhal, A., Yang, P. L., Nibert, M. L., Fichorova, R. N., Lau, D. T., Henrich, T. J., Kaye, K. M., Schachter, S. C., Kuritzkes, D. R., Steinmetz, L. M., Gambhir, S. S., Davis, R. W., Demirci, U. 2015; 112 (32): E4354-63

  Abstract

  Recent advances in biosensing technologies present great potential for medical diagnostics, thus improving clinical decisions. However, creating a label-free general sensing platform capable of detecting multiple biotargets in various clinical specimens over a wide dynamic range, without lengthy sample-processing steps, remains a considerable challenge. In practice, these barriers prevent broad applications in clinics and at patients' homes. Here, we demonstrate the nanoplasmonic electrical field-enhanced resonating device (NE(2)RD), which addresses all these impediments on a single platform. The NE(2)RD employs an immunodetection assay to capture biotargets, and precisely measures spectral color changes by their wavelength and extinction intensity shifts in nanoparticles without prior sample labeling or preprocessing. We present through multiple examples, a label-free, quantitative, portable, multitarget platform by rapidly detecting various protein biomarkers, drugs, protein allergens, bacteria, eukaryotic cells, and distinct viruses. The linear dynamic range of NE(2)RD is five orders of magnitude broader than ELISA, with a sensitivity down to 400 fg/mL This range and sensitivity are achieved by self-assembling gold nanoparticles to generate hot spots on a 3D-oriented substrate for ultrasensitive measurements. We demonstrate that this precise platform handles multiple clinical samples such as whole blood, serum, and saliva without sample preprocessing under diverse conditions of temperature, pH, and ionic strength. The NE(2)RD's broad dynamic range, detection limit, and portability integrated with a disposable fluidic chip have broad applications, potentially enabling the transition toward precision medicine at the point-of-care or primary care settings and at patients' homes.

  View details for DOI 10.1073/pnas.1510824112

  View details for PubMedID 26195743

  View details for PubMedCentralID PMC4538635

• Radiation Dosimetry Study of [(89)Zr]rituximab Tracer for Clinical Translation of B cell NHL Imaging using Positron Emission Tomography. Molecular imaging and biology Natarajan, A., Gambhir, S. S. 2015; 17 (4): 539-547

  Abstract

  We evaluated the dosimetry of [(89)Zr]rituximab, an anti-CD20 immunoPET tracer to image B cell non-Hodgkin's lymphoma (NHL) using a humanized transgenic mouse model that expresses human CD20 transgenic mice (huCD20TM).Rituximab was conjugated to desferrioxamine (Df) for radiolabeling of Zirconium-89. [(89)Zr]rituximab (2.8 ± 0.2 MBq) was tail vein-injected into huCD20T mice. Positron emission tomography (PET)/CT imaging was performed on the two groups of mice (blocking = 2 mg/kg pre-dose of rituximab and non-blocking; n = 5) at eight time points (1, 4, 24, 48, 72, 96, 120, and 168 h) post injection.The novel [(89)Zr]rituximab PET tracer had good immunoreactivity, was stable in human serum, and was able to specifically target human CD20 in mice. The human equivalents of highest dose (mean ± SD) organs with and without pre-dose are liver (345 ± 284 μSv/MBq) and spleen (1165 ± 149 μSv/MBq), respectively.Dosimetry of the human patient whole-body dose was found to be 145 MBq per annum, and the patient dose-limiting organ will be the liver (with rituximab pre-dose blocking) and spleen for non-blocking. The [(89)Zr]rituximab (t½ = 78.4 h) imaging of B cell NHL patients could permit the observation of targeting lesions in NHL patients over an extended period due to longer half-life as compared to the [(64)Cu] rituximab (t½ = 12.7 h).

  View details for DOI 10.1007/s11307-014-0810-8

  View details for PubMedID 25500766

  View details for PubMedCentralID PMC4465424

• Gene expression profiling of individual circulating tumor cells from non-small cell lung cancer (NSCLC) patients via integrated nanotechnologies Park, S., Wong, D. J., Ooi, C., Nair, V. S., Vermesh, O., Lee, S., Suh, S., Lee, L. P., Wang, S. X., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2015

  View details for DOI 10.1158/1538-7445.AM2015-LB-280

  View details for Web of Science ID 000371597100372

• Isolation and Characterization of a Monobody with a Fibronectin Domain III Scaffold That Specifically Binds EphA2 PLOS ONE Park, S., Park, S., Kim, D., Pyo, A., Kimura, R. H., Sathirachinda, A., Choy, H. E., Min, J., Gambhir, S. S., Hong, Y. 2015; 10 (7)

  Abstract

  Monobodies are binding scaffold proteins originating from a human fibronectin domain III (Fn3) scaffold that can be easily engineered with specificity and affinity. Human EphA2 (hEphA2) is an early detection marker protein for various tumors including lung, breast, and colon cancer. In this study, we isolated two hEphA2-specific monobodies (E1 and E10) by screening a yeast surface display library. They showed the same amino acid sequence except in the DE loop and had high affinity (~2 nM Kd) against hEphA2. E1 bound only hEphA2 and mEphA2, although it bound hEphA2 with an affinity 2-fold higher than that of mEphA2. However, E10 also bound the mEphA6 and mEphA8 homologs as well as hEphA2 and mEphA2. Thus, E1 but not E10 was highly specific for hEphA2. E1 specifically bound human cells and xenograft tumor tissues expressing hEphA on the cell surface. In vivo optical imaging showed strong targeting of Cy5.5-labeled E1 to mouse tumor tissue induced by PC3 cells, a human prostate cancer cell line that expresses a high level of hEphA2. In conclusion, the highly specific monobody E1 is useful as a hEphA2 probe candidate for in vivo diagnosis and therapy.

  View details for DOI 10.1371/journal.pone.0132976

  View details for Web of Science ID 000358197600194

  View details for PubMedCentralID PMC4503726

• Development and Validation of an Immuno-PET Tracer as a Companion Diagnostic Agent for Antibody-Drug Conjugate Therapy to Target the CA6 Epitope. Radiology Ilovich, O., Natarajan, A., Hori, S., Sathirachinda, A., Kimura, R., Srinivasan, A., Gebauer, M., Kruip, J., Focken, I., Lange, C., Carrez, C., Sassoon, I., Blanc, V., Sarkar, S. K., Gambhir, S. S. 2015; 276 (1): 191-198

  Abstract

  Purpose To develop and compare three copper 64 ((64)Cu)-labeled antibody fragments derived from a CA6-targeting antibody (huDS6) as immuno-positron emission tomography (immuno-PET)-based companion diagnostic agents for an antibody-drug conjugate by using huDS6. Materials and Methods Three antibody fragments derived from huDS6 were produced, purified, conjugated to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), and evaluated in the following ways: (a) the affinity of the fragments and the DOTA conjugates was measured via flow cytometry, (b) the stability of the labeled fragments was determined ex vivo in human serum over 24 hours, and (c) comparison of the in vivo imaging potential of the fragments was evaluated in mice bearing subcutaneous CA6-positive and CA6-negative xenografts by using serial PET imaging and biodistribution. Isotype controls with antilysozyme and anti-DM4 B-Fabs and blocking experiments with an excess of either B-Fab or huDS6 were used to determine the extent of the antibody fragment (64)Cu-DOTA-B-Fab binding specificity. Immunoreactivity and tracer kinetics were evaluated by using cellular uptake and 48-hour imaging experiments, respectively. Statistical analyses were performed by using t tests, one-way analysis of variance, and Wilcoxon and Mann-Whitney tests. Results The antibody fragment (64)Cu-DOTA-B-Fab was more than 95% stable after 24 hours in human serum, had an immunoreactivity of more than 70%, and allowed differentiation between CA6-positive and CA6-negative tumors in vivo as early as 6 hours after injection, with a 1.7-fold uptake ratio between tumors. Isotype and blocking studies experiments showed tracer-specific uptake in antigen-positive tumors, despite some nonspecific uptake in both tumor models. Conclusion Three antibody fragments were produced and examined as potential companion diagnostic agents. (64)Cu-DOTA-B-Fab is a stable and effective immuno-PET tracer for CA6 imaging in vivo. (©) RSNA, 2015 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.15140058

  View details for PubMedID 25734548

• Early detection of sporadic pancreatic cancer: summative review. Pancreas Chari, S. T., Kelly, K., Hollingsworth, M. A., Thayer, S. P., Ahlquist, D. A., Andersen, D. K., Batra, S. K., Brentnall, T. A., Canto, M., Cleeter, D. F., Firpo, M. A., Gambhir, S. S., Go, V. L., Hines, O. J., Kenner, B. J., Klimstra, D. S., Lerch, M. M., Levy, M. J., Maitra, A., Mulvihill, S. J., Petersen, G. M., Rhim, A. D., Simeone, D. M., Srivastava, S., Tanaka, M., Vinik, A. I., Wong, D. 2015; 44 (5): 693-712

  Abstract

  Pancreatic cancer (PC) is estimated to become the second leading cause of cancer death in the United States by 2020. Early detection is the key to improving survival in PC. Addressing this urgent need, the Kenner Family Research Fund conducted the inaugural Early Detection of Sporadic Pancreatic Cancer Summit Conference in 2014 in conjunction with the 45th Anniversary Meeting of the American Pancreatic Association and Japan Pancreas Society. This seminal convening of international representatives from science, practice, and clinical research was designed to facilitate challenging interdisciplinary conversations to generate innovative ideas leading to the creation of a defined collaborative strategic pathway for the future of the field. An in-depth summary of current efforts in the field, analysis of gaps in specific areas of expertise, and challenges that exist in early detection is presented within distinct areas of inquiry: Case for Early Detection: Definitions, Detection, Survival, and Challenges; Biomarkers for Early Detection; Imaging; and Collaborative Studies. In addition, an overview of efforts in familial PC is presented in an addendum to this article. It is clear from the summit deliberations that only strategically designed collaboration among investigators, institutions, and funders will lead to significant progress in early detection of sporadic PC.

  View details for DOI 10.1097/MPA.0000000000000368

  View details for PubMedID 25931254

  View details for PubMedCentralID PMC4467589

• Early Detection of Sporadic Pancreatic Cancer Summative Review PANCREAS Chari, S. T., Kelly, K., Hollingsworth, M. A., Thayer, S. P., Ahlquist, D. A., Andersen, D. K., Batra, S. K., Brentnall, T. A., Canto, M., Cleeter, D. F., Firpo, M. A., Gambhir, S. S., Go, V. L., Hines, O. J., Kenner, B. J., Klimstra, D. S., Lerch, M. M., Levy, M. J., Maitra, A., Mulvihill, S. J., Petersen, G. M., Rhim, A. D., Simeone, D. M., Srivastava, S., Tanaka, M., Vinik, A. I., Wong, D. 2015; 44 (5): 693-712

  Abstract

  Pancreatic cancer (PC) is estimated to become the second leading cause of cancer death in the United States by 2020. Early detection is the key to improving survival in PC. Addressing this urgent need, the Kenner Family Research Fund conducted the inaugural Early Detection of Sporadic Pancreatic Cancer Summit Conference in 2014 in conjunction with the 45th Anniversary Meeting of the American Pancreatic Association and Japan Pancreas Society. This seminal convening of international representatives from science, practice, and clinical research was designed to facilitate challenging interdisciplinary conversations to generate innovative ideas leading to the creation of a defined collaborative strategic pathway for the future of the field. An in-depth summary of current efforts in the field, analysis of gaps in specific areas of expertise, and challenges that exist in early detection is presented within distinct areas of inquiry: Case for Early Detection: Definitions, Detection, Survival, and Challenges; Biomarkers for Early Detection; Imaging; and Collaborative Studies. In addition, an overview of efforts in familial PC is presented in an addendum to this article. It is clear from the summit deliberations that only strategically designed collaboration among investigators, institutions, and funders will lead to significant progress in early detection of sporadic PC.

  View details for DOI 10.1097/MPA.0000000000000368

  View details for Web of Science ID 000360629300003

  View details for PubMedCentralID PMC4467589

• Predictive Modeling of Drug Response in Non-Hodgkin's Lymphoma PLOS ONE Frieboes, H. B., Smith, B. R., Wang, Z., Kotsuma, M., Ito, K., Day, A., Cahill, B., Flinders, C., Mumenthaler, S. M., Mallick, P., Simbawa, E., Al-Fhaid, A. S., Mahmoud, S. R., Gambhir, S. S., Cristini, V. 2015; 10 (6)

  Abstract

  We combine mathematical modeling with experiments in living mice to quantify the relative roles of intrinsic cellular vs. tissue-scale physiological contributors to chemotherapy drug resistance, which are difficult to understand solely through experimentation. Experiments in cell culture and in mice with drug-sensitive (Eµ-myc/Arf-/-) and drug-resistant (Eµ-myc/p53-/-) lymphoma cell lines were conducted to calibrate and validate a mechanistic mathematical model. Inputs to inform the model include tumor drug transport characteristics, such as blood volume fraction, average geometric mean blood vessel radius, drug diffusion penetration distance, and drug response in cell culture. Model results show that the drug response in mice, represented by the fraction of dead tumor volume, can be reliably predicted from these inputs. Hence, a proof-of-principle for predictive quantification of lymphoma drug therapy was established based on both cellular and tissue-scale physiological contributions. We further demonstrate that, if the in vitro cytotoxic response of a specific cancer cell line under chemotherapy is known, the model is then able to predict the treatment efficacy in vivo. Lastly, tissue blood volume fraction was determined to be the most sensitive model parameter and a primary contributor to drug resistance.

  View details for DOI 10.1371/journal.pone.0129433

  View details for Web of Science ID 000355979500143

  View details for PubMedID 26061425

  View details for PubMedCentralID PMC4464754

• Cu-64-Labeled Divalent Cystine Knot Peptide for Imaging Carotid Atherosclerotic Plaques JOURNAL OF NUCLEAR MEDICINE Jiang, L., Tu, Y., Kimura, R. H., Habte, F., Chen, H., Cheng, K., Shi, H., Gambhir, S. S., Cheng, Z. 2015; 56 (6): 939-944

  Abstract

  The rupture of vulnerable atherosclerotic plaques that lead to stroke and myocardial infarction may be induced by macrophage infiltration and augmented by the expression of integrin αvβ3. Indeed, atherosclerotic angiogenesis may be a promising marker of inflammation. In this study, an engineered integrin αvβ3-targeting PET probe, (64)Cu-NOTA-3-4A, derived from a divalent knottin miniprotein was evaluated in a mouse model for carotid atherosclerotic plaques.Atherosclerotic plaques in BALB/C mice, maintained on a high-fat diet, were induced with streptozotocin injection and carotid artery ligation and verified by MR imaging. Knottin 3-4A was synthesized by solid-phase peptide synthesis chemistry and coupled to 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) before radiolabeling with (64)Cu. PET probe stability in mouse serum was evaluated. Mice with carotid atherosclerotic plaques were injected via the tail vein with (64)Cu-NOTA-3-4A or (18)F-FDG, followed by small-animal PET/CT imaging at different time points. Receptor targeting specificity of the probe was verified by coinjection of c(RGDyK) administered in molar excess. Subsequently, carotid artery dissection and immunofluorescence staining were performed to evaluate target expression.(64)Cu-NOTA-3-4A was synthesized in high radiochemical purity and yield and demonstrated molecular stability in both phosphate-buffered saline and mouse serum at 4 h. Small-animal PET/CT showed that (64)Cu-NOTA-3-4A accumulated at significantly higher levels in the neovasculature of carotid atherosclerotic plaques (7.41 ± 1.44 vs. 0.67 ± 0.23 percentage injected dose/gram, P < 0.05) than healthy or normal vessels at 1 h after injection. (18)F-FDG also accumulated in atherosclerotic lesions at 0.5 and 1 h after injection but at lower plaque-to-normal tissue ratios than (64)Cu-NOTA-3-4A. For example, plaque-to-normal carotid artery ratios for (18)F-FDG and (64)Cu-NOTA-3-4A at 1 h after injection were 3.75 and 14.71 (P < 0.05), respectively. Furthermore, uptake of (64)Cu-NOTA-3-4A in atherosclerotic plaques was effectively blocked (∼90% at 1 h after injection) by coinjection of c(RGDyK). Immunostaining confirmed integrin αvβ3 expression in both the infiltrating macrophages and the neovasculature of atherosclerotic plaques.(64)Cu-NOTA-3-4A demonstrates specific accumulation in carotid atherosclerotic plaques in which macrophage infiltration and angiogenesis are responsible for elevated integrin αvβ3 levels. Therefore, (64)Cu-NOTA-3-4A may demonstrate clinical utility as a PET probe for atherosclerosis imaging or for the evaluation of therapies used to treat atherosclerosis.

  View details for DOI 10.2967/jnumed.115.155176

  View details for Web of Science ID 000355570300026

  View details for PubMedID 25908832

• Development of a High-Throughput Molecular Imaging-Based Orthotopic Hepatocellular Carcinoma Model CUREUS Hwang, G. L., van den Bosch, M. A., Kim, Y. I., Katzenberg, R., Willmann, J. K., Paulmurugan, R., Gambhir, S. S., Hofmann, L. 2015; 7 (6)

  View details for DOI 10.7759/cureus.281

  View details for Web of Science ID 000453603500009

• Development of a High-Throughput Molecular Imaging-Based Orthotopic Hepatocellular Carcinoma Model. Cureus Hwang, G. L., van den Bosch, M. A., Kim, Y. I., Katzenberg, R., Willmann, J. K., Paulmurugan, R., Gambhir, S. S., Hofmann, L. 2015; 7 (6)

  Abstract

  We have developed a novel orthotopic rat hepatocellular (HCC) model and have assessed the ability to use bioluminescence imaging (BLI), positron emission tomography (PET), and ultrasound for early tumor detection and monitoring of disease progression.  Briefly, rat HCC cells were stably transfected with click beetle red as a reporter gene for BLI. Tumor cells were injected under direct visualization into the left or middle lobe of the liver in 37 rats. In six animals, serial PET, BLI, and ultrasound imaging were performed at 10-time points in 28 days. The remainder of the animals underwent PET imaging at 14 days. Tumor implantation was successful in 34 of 37 animals (91.9%). In the six animals that underwent serial imaging, tumor formation was first detected with BLI on Day 4 with continued increase through Day 21, and hypermetabolic activity on PET was first noted on Days 14-15 with continued increase through Day 28. PET activity was seen on Day 14 in the 28 other animals that demonstrated tumor development. Anatomic tumor formation was detected with ultrasound at Days 10-12 with continued growth through Day 28. The first metastases were detected by PET after Day 24.        We have successfully developed and validated a novel orthotopic HCC small animal model that permits longitudinal assessment of change in tumor size using molecular imaging techniques. BLI is the most sensitive imaging method for detection of early tumor formation and growth. This model permits high-throughput in vivo evaluation of image-guided therapies.

  View details for DOI 10.7759/cureus.281

  View details for PubMedID 26180705

  View details for PubMedCentralID PMC4494575

• Glioblastoma Multiforme Recurrence: An Exploratory Study of (18)F FPPRGD2 PET/CT. Radiology Iagaru, A., Mosci, C., Mittra, E., Zaharchuk, G., Fischbein, N., Harsh, G., Li, G., Nagpal, S., Recht, L., Gambhir, S. S. 2015: 141550

  Abstract

  Purpose To prospectively evaluate fluorine 18 ((18)F) 2-fluoropropionyl-labeled PEGylated dimeric arginine-glycine-aspartic acid (RGD) peptide (PEG3-E[c{RGDyk}]2) (FPPRGD2) positron emission tomography (PET) in patients with glioblastoma multiforme (GBM). Materials and Methods The institutional review board approved this HIPAA-compliant protocol. Written informed consent was obtained from each patient. (18)F FPPRGD2 uptake was measured semiquantitatively in the form of maximum standardized uptake values (SUVmax) and uptake volumes before and after treatment with bevacizumab. Vital signs and laboratory results were collected before, during, and after the examinations. A nonparametric version of multivariate analysis of variance was used to assess safety outcome measures simultaneously across time points. A paired two-sample t test was performed to compare SUVmax. Results A total of 17 participants (eight men, nine women; age range, 25-65 years) were enrolled prospectively. (18)F FPPRGD2 PET/computed tomography (CT), (18)F fluorodeoxyglucose (FDG) PET/CT, and brain magnetic resonance (MR) imaging were performed within 3 weeks, prior to the start of bevacizumab therapy. In eight of the 17 patients (47%), (18)F FPPRGD2 PET/CT was repeated 1 week after the start of bevacizumab therapy; six patients (35%) underwent (18)F FPPRGD2 PET/CT a third time 6 weeks after starting bevacizumab therapy. There were no changes in vital signs, electrocardiographic findings, or laboratory values that qualified as adverse events. One patient (6%) had recurrent GBM identified only on (18)F FPPRGD2 PET images, and subsequent MR images enabled confirmation of recurrence. Of the 17 patients, 14 (82%) had recurrent GBM identified on (18)F FPPRGD2 PET and brain MR images, while (18)F FDG PET enabled identification of recurrence in 13 (76%) patients. Two patients (12%) had no recurrent GBM. Conclusion (18)F FPPRGD2 is a safe PET radiopharmaceutical that has increased uptake in GBM lesions. Larger cohorts are required to confirm these preliminary findings. (©) RSNA, 2015 Online supplemental material is available for this article.

  View details for DOI 10.1148/radiol.2015141550

  View details for PubMedID 25965900

• Semiquantitative Analysis of the Biodistribution of the Combined F-18-NaF and F-18-FDG Administration for PET/CT Imaging JOURNAL OF NUCLEAR MEDICINE Minamimoto, R., Mosci, C., Jamali, M., Barkhodari, A., Habte, F., Jackson, T., Mittra, E., Gambhir, S. S., Iagaru, A. 2015; 56 (5): 688-694

  Abstract

  In this study we evaluated the biodistribution of the (18)F-/(18)F-FDG administration compared to separate (18)F-NaF and (18)F-FDG. We also estimated the interaction of (18)F-NaF and (18)F-FDG in the (18)F-/(18)F-FDG administration by semiquantitative analysis.We retrospectively analyzed data of 49 patients (male 39, female 10; mean ± SD age: 59.3 ± 15.2 years) who had separate (18)F-FDG PET/CT and (18)F-NaF PET/CT, as well as the (18)F-/(18)F-FDG PET/CT sequentially. The most common primary diagnosis was prostate cancer (n = 28), followed by sarcoma (n = 9) and breast cancer (n = 6). The mean standardized uptake values (SUVmean) were recorded for 18 organs in all patients, while maximum SUV (SUVmax) and SUVmean were recorded for all the identified malignant lesions. We also estimated the (18)F-/(18)F-FDG uptake by sum of (18)F-FDG uptake and adjusted (18)F-NaF uptake based on the ratio of (18)F-NaF injected dose in (18)F-/(18)F-FDG PET/CT. Lastly, we compared the results in order to explore the interaction of (18)F-FDG and (18)F-NaF uptake in the (18)F-/(18)F-FDG scan.The (18)F-/(18)F-FDG uptake in the cerebral cortex, cerebellum, parotid grand, myocardium and bowel mostly reflect the (18)F-FDG uptake, while the uptake in the other analyzed structures is influenced by both the (18)F-FDG and the (18)F-NaF uptake. The (18)F-/(18)F-FDG uptake in extra skeletal lesions shows no significant difference when compared to the uptake from the separate (18)F-FDG scan. The (18)F-/(18)F-FDG uptake in skeletal lesions reflected mostly the (18)F-NaF uptake. Tumor to background (T/B) ratio of (18)F-/(18)F-FDG in extra skeletal lesions showed no significant difference when compared with that from (18)F-FDG alone (P = 0.73). For skeletal lesions, T/B ratio of (18)F-/(18)F-FDG was lower than that from (18)F-NaF alone (P <0.001); however, this difference did not result in missed skeletal lesions on the (18)F-/(18)F-FDG scan.The understanding of the biodistribution of radiopharmaceuticals and the lesions uptake of the (18)F-/(18)F-FDG scan, as well as the variations compared to the uptake on the separate (18)F-FDG PET/CT and (18)F-NaF PET/CT are valuable for more in depth evaluation of the combined scanning technique.

  View details for DOI 10.2967/jnumed.115.153767

  View details for Web of Science ID 000353831000013

  View details for PubMedID 25840978

• Imaging of tumor-associated system x(C)(-) activity with 18F-fluoropropylglutamate (FSPG) PET/CT for intracranial malignancies. Mittra, E., Minamimoto, R., Barkhodari, A., Jamali, M., Schneider, B., Koglin, N., Berndt, M., Stephens, A., Chin, F., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738803222

• Imaging poly(ADP ribose) polymerase-1 activity for personalized cancer medicine using a novel PET tracer Shuhendler, A., Cui, L., Lin, J., Shen, B., James, M., Witney, T., Chattopadhyay, N., Gambhir, S., Chin, F., Rao, J. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738800003

• PET Imaging Carotid Atherosclerostic Plaque Using Divalent Knottin Jiang, L., Tu, Y., Kimura, R., Habte, F., Chen, H., Cheng, K., Shi, H., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738801112

• F-18 FPPRGD(2) PET as a Surrogate Biomarker of Integrin alpha(v)beta(3) Expression Before and After Anti-angiogenesis Treatment 18 Minamimoto, R., Jamali, M., Barkhodari, A., Mosci, C., Mittra, E., Shen, B., Chin, F., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738801280

• Antibody mimics, fibronectin domain III for EphA2-targeting as a probe in murine tumor model Park, S., Hong, Y., Park, S., Kimura, R., Min, J., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738802114

• Physiological distribution of Ga-68-DOTA-TATE: an atlas of standardized uptake values Moradi, F., Minamimoto, R., Jamali, M., Barkhodari, A., Quon, A., Mittra, E., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738802255

• Prospective evaluation of Tc-99m MDP scintigraphy, F-18 NaF/F-18 FDG PET/CT and WBMRI in patients with breast and prostate cancers Iagaru, A., Minamimoto, R., Mosci, C., Jamali, M., Barkhodari, A., Loening, A., Taviani, V., Mittra, E., Gambhir, S., Vasanawala, S. SOC NUCLEAR MEDICINE INC. 2015

  View details for Web of Science ID 000358738802275

• Optical coherence contrast imaging using gold nanorods in living mice eyes CLINICAL AND EXPERIMENTAL OPHTHALMOLOGY de la Zerda, A., Prabhulkar, S., Perez, V. L., Ruggeri, M., Paranjape, A. S., Habte, F., Gambhir, S. S., Awdeh, R. M. 2015; 43 (4): 358-366

  Abstract

  Optical coherence tomography (OCT) is a powerful imaging modality to visualize tissue structures, with axial image pixel resolution as high as 1.6 μm in tissue. However, OCT is intrinsically limited to providing structural information as the OCT contrast is produced by optically scattering tissues.Gold nanorods (GNRs) were injected into the anterior chamber (AC) and cornea of mice eyes which could create a significant OCT signal and hence could be used as a contrast agent for in vivo OCT imaging.A dose of 30 nM of GNRs (13 nm in diameter and 45 nm in length) were injected to the AC of mice eyes and produced an OCT contrast nearly 50-fold higher than control mice injected with saline. Furthermore, the lowest detectable concentration of GNRs in living mice AC was experimentally estimated to be as low as 120 pM.The high sensitivity and low toxicity of GNRs brings great promise for OCT to uniquely become a high-resolution molecular imaging modality.

  View details for DOI 10.1111/ceo.12299

  View details for Web of Science ID 000356810200009

  View details for PubMedID 24533647

• A Real-Time Clinical Endoscopic System for Intraluminal, Multiplexed Imaging of Surface-Enhanced Raman Scattering Nanoparticles PLOS ONE Garai, E., Sensarn, S., Zavaleta, C. L., Loewke, N. O., Rogalla, S., Mandella, M. J., Felt, S. A., Friedland, S., Liu, J. T., Gambhir, S. S., Contag, C. H. 2015; 10 (4)

  Abstract

  The detection of biomarker-targeting surface-enhanced Raman scattering (SERS) nanoparticles (NPs) in the human gastrointestinal tract has the potential to improve early cancer detection; however, a clinically relevant device with rapid Raman-imaging capability has not been described. Here we report the design and in vivo demonstration of a miniature, non-contact, opto-electro-mechanical Raman device as an accessory to clinical endoscopes that can provide multiplexed molecular data via a panel of SERS NPs. This device enables rapid circumferential scanning of topologically complex luminal surfaces of hollow organs (e.g., colon and esophagus) and produces quantitative images of the relative concentrations of SERS NPs that are present. Human and swine studies have demonstrated the speed and simplicity of this technique. This approach also offers unparalleled multiplexing capabilities by simultaneously detecting the unique spectral fingerprints of multiple SERS NPs. Therefore, this new screening strategy has the potential to improve diagnosis and to guide therapy by enabling sensitive quantitative molecular detection of small and otherwise hard-to-detect lesions in the context of white-light endoscopy.

  View details for DOI 10.1371/journal.pone.0123185

  View details for Web of Science ID 000353711600032

  View details for PubMedID 25923788

  View details for PubMedCentralID PMC4414592

• Synthesis of [(18)F]-labelled Maltose Derivatives as PET Tracers for Imaging Bacterial Infection. Molecular imaging and biology Namavari, M., Gowrishankar, G., Hoehne, A., Jouannot, E., Gambhir, S. S. 2015; 17 (2): 168-176

  Abstract

  To develop novel positron emission tomography (PET) agents for visualization and therapy monitoring of bacterial infections.It is known that maltose and maltodextrins are energy sources for bacteria. Hence, (18)F-labelled maltose derivatives could be a valuable tool for imaging bacterial infections. We have developed methods to synthesize 4-O-(α-D-glucopyranosyl)-6-deoxy-6-[(18)F]fluoro-D-glucopyranoside (6-[(18)F]fluoromaltose) and 4-O-(α-D-glucopyranosyl)-1-deoxy-1-[(18)F]fluoro-D-glucopyranoside (1-[(18)F]fluoromaltose) as bacterial infection PET imaging agents. 6-[(18)F]fluoromaltose was prepared from precursor 1,2,3-tri-O-acetyl-4-O-(2',3',-di-O-acetyl-4',6'-benzylidene-α-D-glucopyranosyl)-6-deoxy-6-nosyl-D-glucopranoside (5). The synthesis involved the radio-fluorination of 5 followed by acidic and basic hydrolysis to give 6-[(18)F]fluoromaltose. In an analogous procedure, 1-[(18)F]fluoromaltose was synthesized from 2,3, 6-tri-O-acetyl-4-O-(2',3',4',6-tetra-O-acetyl-α-D-glucopyranosyl)-1-deoxy-1-O-triflyl-D-glucopranoside (9). Stability of 6-[(18)F]fluoromaltose in phosphate-buffered saline (PBS) and human and mouse serum at 37 °C was determined. Escherichia coli uptake of 6-[(18)F]fluoromaltose was examined.A reliable synthesis of 1- and 6-[(18)F]fluoromaltose has been accomplished with 4-6 and 5-8 % radiochemical yields, respectively (decay-corrected with 95 % radiochemical purity). 6-[(18)F]fluoromaltose was sufficiently stable over the time span needed for PET studies (∼96 % intact compound after 1-h and ∼65 % after 2-h incubation in serum). Bacterial uptake experiments indicated that E. coli transports 6-[(18)F]fluoromaltose. Competition assays showed that the uptake of 6-[(18)F]fluoromaltose was completely blocked by co-incubation with 1 mM of the natural substrate maltose.We have successfully synthesized 1- and 6-[(18)F]fluoromaltose via direct fluorination of appropriate protected maltose precursors. Bacterial uptake experiments in E. coli and stability studies suggest a possible application of 6-[(18)F]fluoromaltose as a new PET imaging agent for visualization and monitoring of bacterial infections.

  View details for DOI 10.1007/s11307-014-0793-5

  View details for PubMedID 25277604

• Detecting cancers through tumor-activatable minicircles that lead to a detectable blood biomarker. Proceedings of the National Academy of Sciences of the United States of America Ronald, J. A., Chuang, H., Dragulescu-Andrasi, A., Hori, S. S., Gambhir, S. S. 2015; 112 (10): 3068-3073

  Abstract

  Earlier detection of cancers can dramatically improve the efficacy of available treatment strategies. However, despite decades of effort on blood-based biomarker cancer detection, many promising endogenous biomarkers have failed clinically because of intractable problems such as highly variable background expression from nonmalignant tissues and tumor heterogeneity. In this work we present a tumor-detection strategy based on systemic administration of tumor-activatable minicircles that use the pan-tumor-specific Survivin promoter to drive expression of a secretable reporter that is detectable in the blood nearly exclusively in tumor-bearing subjects. After systemic administration we demonstrate a robust ability to differentiate mice bearing human melanoma metastases from tumor-free subjects for up to 2 wk simply by measuring blood reporter levels. Cumulative change in reporter levels also identified tumor-bearing subjects, and a receiver operator-characteristic curve analysis highlighted this test's performance with an area of 0.918 ± 0.084. Lung tumor burden additionally correlated (r(2) = 0.714; P < 0.05) with cumulative reporter levels, indicating that determination of disease extent was possible. Continued development of our system could improve tumor detectability dramatically because of the temporally controlled, high reporter expression in tumors and nearly zero background from healthy tissues. Our strategy's highly modular nature also allows it to be iteratively optimized over time to improve the test's sensitivity and specificity. We envision this system could be used first in patients at high risk for tumor recurrence, followed by screening high-risk populations before tumor diagnosis, and, if proven safe and effective, eventually may have potential as a powerful cancer-screening tool for the general population.

  View details for DOI 10.1073/pnas.1414156112

  View details for PubMedID 25713388

  View details for PubMedCentralID PMC4364239

• Detection of osseous metastasis by 18F-NaF/18F-FDG PET/CT versus CT alone. Clinical nuclear medicine Sampath, S. C., Sampath, S. C., Mosci, C., Lutz, A. M., Willmann, J. K., Mittra, E. S., Gambhir, S. S., Iagaru, A. 2015; 40 (3): e173-7

  Abstract

  Sodium fluoride PET (F-NaF) has recently reemerged as a valuable method for detection of osseous metastasis, with recent work highlighting the potential of coadministered F-NaF and F-FDG PET/CT in a single combined imaging examination. We further examined the potential of such combined examinations by comparing dual tracer F-NaF/F-FDG PET/CT with CT alone for detection of osseous metastasis.Seventy-five participants with biopsy-proven malignancy were consecutively enrolled from a single center and underwent combined F-NaF/F-FDG PET/CT and diagnostic CT scans. PET/CT as well as CT only images were reviewed in blinded fashion and compared with the results of clinical, imaging, or histological follow-up as a truth standard.Sensitivity of the combined F-NaF/F-FDG PET/CT was higher than that of CT alone (97.4% vs 66.7%). CT and F-NaF/F-FDG PET/CT were concordant in 73% of studies. Of 20 discordant cases, F-NaF/F-FDG PET/CT was correct in 19 (95%). Three cases were interpreted concordantly but incorrectly, and all 3 were false positives. A single case of osseous metastasis was detected by CT alone, but not by F-NaF/F-FDG PET/CT.Combined F-NaF/F-FDG PET/CT outperforms CT alone and is highly sensitive and specific for detection of osseous metastases. The concordantly interpreted false-positive cases demonstrate the difficulty of distinguishing degenerative from malignant disease, whereas the single case of metastasis seen on CT but not PET highlights the need for careful review of CT images in multimodality studies.

  View details for DOI 10.1097/RLU.0000000000000560

  View details for PubMedID 25140557

• 18F-FPRGD2 PET/CT imaging of integrin avß3 in renal carcinomas: correlation with histopathology. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Withofs, N., Signolle, N., Somja, J., Lovinfosse, P., Nzaramba, E. M., Mievis, F., Giacomelli, F., Waltregny, D., Cataldo, D., Gambhir, S. S., Hustinx, R. 2015; 56 (3): 361-364

  Abstract

  This study aimed to correlate (18)F-FB-mini-PEG-E[c(RGDyK)](2) ((18)F-FPRGD2) uptake to integrin αvβ3 expression and angiogenesis in renal tumors.(18)F-FPRGD2 PET/CT was performed on 27 patients before surgical resection (median 4 d) of a renal mass. The (18)F-FPRGD2 uptake was compared with integrin αvβ3, CD31, CD105, and Ki-67 using immunohistochemistry; with placental growth factor and vascular endothelial growth factor receptors 1 and 2 using reverse transcription polymerase chain reaction; and with vascular endothelial growth factor A isoforms using enzyme-linked immunosorbent assay.Overall, (18)F-FPRGD2 uptake significantly correlated (P < 0.0001) with integrin αvβ3 expression in renal masses. However, it correlated only with integrin αvβ3-positive vessels in the group of papillary carcinomas whereas it correlated with integrin αvβ3 expression by tumor cells in the clear cell carcinoma group.(18)F-FPRGD2 uptake reflects the expression of integrin αvβ3 in renal tumors but represents angiogenesis only when tumor cells do not express the integrin.

  View details for DOI 10.2967/jnumed.114.149021

  View details for PubMedID 25655629

• F-18-FPRGD2 PET/CT Imaging of Integrin alpha(v)beta(3) in Renal Carcinomas: Correlation with Histopathology JOURNAL OF NUCLEAR MEDICINE Withofs, N., Signolle, N., Somja, J., Lovinfosse, P., Nzaramba, E. M., Mievis, F., Giacomelli, F., Waltregny, D., Cataldo, D., Gambhir, S. S., Hustinx, R. 2015; 56 (3): 361-364

  Abstract

  This study aimed to correlate (18)F-FB-mini-PEG-E[c(RGDyK)](2) ((18)F-FPRGD2) uptake to integrin αvβ3 expression and angiogenesis in renal tumors.(18)F-FPRGD2 PET/CT was performed on 27 patients before surgical resection (median 4 d) of a renal mass. The (18)F-FPRGD2 uptake was compared with integrin αvβ3, CD31, CD105, and Ki-67 using immunohistochemistry; with placental growth factor and vascular endothelial growth factor receptors 1 and 2 using reverse transcription polymerase chain reaction; and with vascular endothelial growth factor A isoforms using enzyme-linked immunosorbent assay.Overall, (18)F-FPRGD2 uptake significantly correlated (P < 0.0001) with integrin αvβ3 expression in renal masses. However, it correlated only with integrin αvβ3-positive vessels in the group of papillary carcinomas whereas it correlated with integrin αvβ3 expression by tumor cells in the clear cell carcinoma group.(18)F-FPRGD2 uptake reflects the expression of integrin αvβ3 in renal tumors but represents angiogenesis only when tumor cells do not express the integrin.

  View details for DOI 10.2967/jnumed.114.149021

  View details for Web of Science ID 000350631700032

• Detection of Osseous Metastasis by 18F-NaF/18F-FDG PET/CT Versus CT Alone. Clinical nuclear medicine Sampath, S. C., Sampath, S. C., Mosci, C., Lutz, A. M., Willmann, J. K., Mittra, E. S., Gambhir, S. S., Iagaru, A. 2015; 40 (3): e173-7

  Abstract

  Sodium fluoride PET (F-NaF) has recently reemerged as a valuable method for detection of osseous metastasis, with recent work highlighting the potential of coadministered F-NaF and F-FDG PET/CT in a single combined imaging examination. We further examined the potential of such combined examinations by comparing dual tracer F-NaF/F-FDG PET/CT with CT alone for detection of osseous metastasis.Seventy-five participants with biopsy-proven malignancy were consecutively enrolled from a single center and underwent combined F-NaF/F-FDG PET/CT and diagnostic CT scans. PET/CT as well as CT only images were reviewed in blinded fashion and compared with the results of clinical, imaging, or histological follow-up as a truth standard.Sensitivity of the combined F-NaF/F-FDG PET/CT was higher than that of CT alone (97.4% vs 66.7%). CT and F-NaF/F-FDG PET/CT were concordant in 73% of studies. Of 20 discordant cases, F-NaF/F-FDG PET/CT was correct in 19 (95%). Three cases were interpreted concordantly but incorrectly, and all 3 were false positives. A single case of osseous metastasis was detected by CT alone, but not by F-NaF/F-FDG PET/CT.Combined F-NaF/F-FDG PET/CT outperforms CT alone and is highly sensitive and specific for detection of osseous metastases. The concordantly interpreted false-positive cases demonstrate the difficulty of distinguishing degenerative from malignant disease, whereas the single case of metastasis seen on CT but not PET highlights the need for careful review of CT images in multimodality studies.

  View details for DOI 10.1097/RLU.0000000000000560

  View details for PubMedID 25140557

• PET Imaging of Translocator Protein (18 kDa) in a Mouse Model of Alzheimer's Disease Using N-(2,5-Dimethoxybenzyl)-2-18F-Fluoro-N-(2-Phenoxyphenyl)Acetamide. Journal of nuclear medicine : official publication, Society of Nuclear Medicine James, M. L., Belichenko, N. P., Nguyen, T. V., Andrews, L. E., Ding, Z., Liu, H., Bodapati, D., Arksey, N., Shen, B., Cheng, Z., Wyss-Coray, T., Gambhir, S. S., Longo, F. M., Chin, F. T. 2015; 56 (2): 311-316

  Abstract

  Herein we aimed to evaluate the utility of N-(2,5-dimethoxybenzyl)-2-(18)F-fluoro-N-(2-phenoxyphenyl)acetamide ((18)F-PBR06) for detecting alterations in translocator protein (TSPO) (18 kDa), a biomarker of microglial activation, in a mouse model of Alzheimer's disease (AD).Wild-type (wt) and AD mice (i.e., APP(L/S)) underwent (18)F-PBR06 PET imaging at predetermined time points between the ages of 5-6 and 15-16 mo. MR images were fused with PET/CT data to quantify (18)F-PBR06 uptake in the hippocampus and cortex. Ex vivo autoradiography and TSPO/CD68 immunostaining were also performed using brain tissue from these mice.PET images showed significantly higher accumulation of (18)F-PBR06 in the cortex and hippocampus of 15- to 16-mo-old APP(L/S) mice than age-matched wts (cortex/muscle: 2.43 ± 0.19 vs. 1.55 ± 0.15, P < 0.005; hippocampus/muscle: 2.41 ± 0.13 vs. 1.55 ± 0.12, P < 0.005). And although no significant difference was found between wt and APP(L/S) mice aged 9-10 mo or less using PET (P = 0.64), we were able to visualize and quantify a significant difference in (18)F-PBR06 uptake in these mice using autoradiography (cortex/striatum: 1.13 ± 0.04 vs. 0.96 ± 0.01, P < 0.05; hippocampus/striatum: 1.266 ± 0.003 vs. 1.096 ± 0.017, P < 0.001). PET results for 15- to 16-mo-old mice correlated well with autoradiography and immunostaining (i.e., increased (18)F-PBR06 uptake in brain regions containing elevated CD68 and TSPO staining in APP(L/S) mice, compared with wts).(18)F-PBR06 shows great potential as a tool for visualizing TSPO/microglia in the progression and treatment of AD.

  View details for DOI 10.2967/jnumed.114.141648

  View details for PubMedID 25613536

• Sol-Gel Synthesis and Electrospraying of Biodegradable (P2O5)(55)-(CaO)(30)-(Na2O)(15) Glass Nanospheres as a Transient Contrast Agent for Ultrasound Stem Cell Imaging ACS NANO Foroutan, F., Jokerst, J. V., Gambhir, S. S., Vermesh, O., Kim, H., Knowles, J. C. 2015; 9 (2): 1868-1877

  Abstract

  Ultrasound imaging is a powerful tool in medicine because of the millisecond temporal resolution and submillimeter spatial resolution of acoustic imaging. However, the current generation of acoustic contrast agents is primarily limited to vascular targets due to their large size. Nanosize particles have the potential to be used as a contrast agent for ultrasound molecular imaging. Silica-based nanoparticles have shown promise here; however, their slow degradation rate may limit their applications as a contrast agent. Phosphate-based glasses are an attractive alternative with controllable degradation rate and easily metabolized degradation components in the body. In this study, biodegradable P2O5-CaO-Na2O phosphate-based glass nanospheres (PGNs) were synthesized and characterized as contrast agents for ultrasound imaging. The structure of the PGNs was characterized using scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), (31)P magic angle spinning nuclear magnetic resonance ((31)P MAS NMR), and Fourier transform infrared (FTIR) spectroscopy. The SEM images indicated a spherical shape with a diameter size range of 200-500 nm. The XRD, (31)P NMR, and FTIR results revealed the amorphous and glassy nature of PGNs that consisted of mainly Q(1) and Q(2) phosphate units. We used this contrast to label mesenchymal stem cells and determined in vitro and in vivo detection limits of 5 and 9 μg/mL, respectively. Cell counts down to 4000 could be measured with ultrasound imaging with no cytoxicity at doses needed for imaging. Importantly, ion-release studies confirmed these PGNs biodegrade into aqueous media with degradation products that can be easily metabolized in the body.

  View details for DOI 10.1021/nn506789y

  View details for PubMedID 25625373

• 18F-FAZA PET Imaging Response Tracks the Reoxygenation of Tumors in Mice upon Treatment with the Mitochondrial Complex I Inhibitor BAY 87-2243. Clinical cancer research Chang, E., Liu, H., Unterschemmann, K., Ellinghaus, P., Liu, S., Gekeler, V., Cheng, Z., Berndorff, D., Gambhir, S. S. 2015; 21 (2): 335-346

  Abstract

  We describe a noninvasive PET imaging method that monitors early therapeutic efficacy of BAY 87-2243, a novel small-molecule inhibitor of mitochondrial complex I as a function of hypoxia-inducible factor-1α (HIF1α) activity.Four PET tracers [(18)F-FDG, (18)F-Fpp(RGD)2, (18)F-FLT, and (18)F-FAZA] were assessed for uptake into tumor xenografts of drug-responsive (H460, PC3) or drug-resistant (786-0) carcinoma cells. Mice were treated with BAY 87-2243 or vehicle. At each point, RNA from treated and vehicle H460 tumor xenografts (n = 3 each) was isolated and analyzed for target genes.Significant changes in uptake of (18)F-FAZA, (18)F-FLT, and (18)F-Fpp(RGD)2 (P < 0.01) occurred with BAY 87-2243 treatment with (18)F-FAZA being the most prominent. (18)F-FDG uptake was unaffected. (18)F-FAZA tumor uptake declined by 55% to 70% (1.21% ± 0.10%ID/g to 0.35 ± 0.1%ID/g; n = 6, vehicle vs. treatment) in both H460 (P < 0.001) and PC3 (P < 0.05) xenografts 1 to 3 days after drug administration. (18)F-FAZA uptake in 786-0 xenografts was unaffected. Decline occurred before significant differences in tumor volume, thus suggesting (18)F-FAZA decrease reflected early changes in tumor metabolism. BAY 87-2243 reduced expression of hypoxia-regulated genes CA IX, ANGPTL4, and EGLN-3 by 99%, 93%, and 83%, respectively (P < 0.001 for all), which corresponds with reduced (18)F-FAZA uptake upon drug treatment. Heterogeneous expression of genes associated with glucose metabolism, vessel density, and proliferation was observed.Our studies suggest suitability of (18)F-FAZA-PET as an early pharmacodynamic monitor on the efficacy of anticancer agents that target the mitochondrial complex I and intratumor oxygen levels (e.g., BAY 87-2243). Clin Cancer Res; 21(2); 335-46. ©2014 AACR.

  View details for DOI 10.1158/1078-0432.CCR-14-0217

  View details for PubMedID 25381339

  View details for PubMedCentralID PMC4297600

• Simultaneous Whole-Body Time-of-Flight F-18-FDG PET/MRI A Pilot Study Comparing SUVmax With PET/CT and Assessment of MR Image Quality CLINICAL NUCLEAR MEDICINE Iagaru, A., Mittra, E., Minamimoto, R., Jamali, M., Levin, C., Quon, A., Gold, G., Herfkens, R., Vasanawala, S., Gambhir, S. S., Zaharchuk, G. 2015; 14 (1): 1-8

  Abstract

  The recent introduction of hybrid PET/MRI scanners in clinical practice has shown promising initial results for several clinical scenarios. However, the first generation of combined PET/MRI lacks time-of-flight (TOF) technology. Here we report the results of the first patients to be scanned on a completely novel fully integrated PET/MRI scanner with TOF.We analyzed data from patients who underwent a clinically indicated F FDG PET/CT, followed by PET/MRI. Maximum standardized uptake values (SUVmax) were measured from F FDG PET/MRI and F FDG PET/CT for lesions, cerebellum, salivary glands, lungs, aortic arch, liver, spleen, skeletal muscle, and fat. Two experienced radiologists independently reviewed the MR data for image quality.Thirty-six patients (19 men, 17 women, mean [±standard deviation] age of 61 ± 14 years [range: 27-86 years]) with a total of 69 discrete lesions met the inclusion criteria. PET/CT images were acquired at a mean (±standard deviation) of 74 ± 14 minutes (range: 49-100 minutes) after injection of 10 ± 1 mCi (range: 8-12 mCi) of F FDG. PET/MRI scans started at 161 ± 29 minutes (range: 117 - 286 minutes) after the F FDG injection. All lesions identified on PET from PET/CT were also seen on PET from PET/MRI. The mean SUVmax values were higher from PET/MRI than PET/CT for all lesions. No degradation of MR image quality was observed.The data obtained so far using this investigational PET/MR system have shown that the TOF PET system is capable of excellent performance during simultaneous PET/MR with routine pulse sequences. MR imaging was not compromised. Comparison of the PET images from PET/CT and PET/MRI show no loss of image quality for the latter. These results support further investigation of this novel fully integrated TOF PET/MRI instrument.

  View details for Web of Science ID 000346633400023

• A Magnetic Bead-Based Sensor for the Quantification of Multiple Prostate Cancer Biomarkers. PloS one Jokerst, J. V., Chen, Z., Xu, L., Nolley, R., Chang, E., Mitchell, B., Brooks, J. D., Gambhir, S. S. 2015; 10 (9): e0139484

  Abstract

  Novel biomarker assays and upgraded analytical tools are urgently needed to accurately discriminate benign prostatic hypertrophy (BPH) from prostate cancer (CaP). To address this unmet clinical need, we report a piezeoelectric/magnetic bead-based assay to quantitate prostate specific antigen (PSA; free and total), prostatic acid phosphatase, carbonic anhydrase 1 (CA1), osteonectin, IL-6 soluble receptor (IL-6sr), and spondin-2. We used the sensor to measure these seven proteins in serum samples from 120 benign prostate hypertrophy patients and 100 Gleason score 6 and 7 CaP using serum samples previously collected and banked. The results were analyzed with receiver operator characteristic curve analysis. There were significant differences between BPH and CaP patients in the PSA, CA1, and spondin-2 assays. The highest AUC discrimination was achieved with a spondin-2 OR free/total PSA operation-the area under the curve was 0.84 with a p value below 10-6. Some of these data seem to contradict previous reports and highlight the importance of sample selection and proper assay building in the development of biomarker measurement schemes. This bead-based system offers important advantages in assay building including low cost, high throughput, and rapid identification of an optimal matched antibody pair.

  View details for DOI 10.1371/journal.pone.0139484

  View details for PubMedID 26421725

  View details for PubMedCentralID PMC4589536

• Validation of 64Cu-DOTA-rituximab injection preparation under good manufacturing practices: a PET tracer for imaging of B-cell non-Hodgkin lymphoma. Molecular imaging Natarajan, A., Arksey, N., Iagaru, A., Chin, F. T., Gambhir, S. S. 2015; 14

  View details for DOI 10.2310/7290.2014.00055

  View details for PubMedID 25762106

• A Magnetic Bead-Based Sensor for the Quantification of Multiple Prostate Cancer Biomarkers. PloS one Jokerst, J. V., Chen, Z., Xu, L., Nolley, R., Chang, E., Mitchell, B., Brooks, J. D., Gambhir, S. S. 2015; 10 (9)

  Abstract

  Novel biomarker assays and upgraded analytical tools are urgently needed to accurately discriminate benign prostatic hypertrophy (BPH) from prostate cancer (CaP). To address this unmet clinical need, we report a piezeoelectric/magnetic bead-based assay to quantitate prostate specific antigen (PSA; free and total), prostatic acid phosphatase, carbonic anhydrase 1 (CA1), osteonectin, IL-6 soluble receptor (IL-6sr), and spondin-2. We used the sensor to measure these seven proteins in serum samples from 120 benign prostate hypertrophy patients and 100 Gleason score 6 and 7 CaP using serum samples previously collected and banked. The results were analyzed with receiver operator characteristic curve analysis. There were significant differences between BPH and CaP patients in the PSA, CA1, and spondin-2 assays. The highest AUC discrimination was achieved with a spondin-2 OR free/total PSA operation-the area under the curve was 0.84 with a p value below 10-6. Some of these data seem to contradict previous reports and highlight the importance of sample selection and proper assay building in the development of biomarker measurement schemes. This bead-based system offers important advantages in assay building including low cost, high throughput, and rapid identification of an optimal matched antibody pair.

  View details for DOI 10.1371/journal.pone.0139484

  View details for PubMedID 26421725

  View details for PubMedCentralID PMC4589536

• Parts per billion detection of uranium with a porphyrinoid-containing nanoparticle and in vivo photoacoustic imaging ANALYST Ho, I., Sessler, J. L., Gambhir, S. S., Jokerst, J. V. 2015; 140 (11): 3731-3737

  Abstract

  Chemical tools that can report radioactive isotopes would be of interest to the defense community. Here we report ∼250 nm polymeric nanoparticles containing porphyrinoid macrocycles with and without pre-complexed depleted uranium and demonstrate that the latter species may be detected easily and with high sensitivity via photoacoustic imaging. The porphyrinoid macrocycles used in the present study are non-aromatic in the absence of the uranyl cation, but aromatic after cation complexation. We solubilized both the freebase and metalated forms of the macrocycles in poly(lactic-co-glycolic acid) and found a peak in the photoacoustic spectrum at 910 nm excitation in the case of the uranyl complex. The signal was stable for at least 15 minutes and allowed detection of uranium concentrations down to 6.2 ppb (5.7 nM) in vitro and 0.57 ppm (19 fCi; 0.52 μM) in vivo. To the best of our knowledge, this is the first report of a nanoparticle that detects an actinide cation via photoacoustic imaging.

  View details for DOI 10.1039/c5an00207a

  View details for PubMedID 25854506

• Theranostic mesoporous silica nanoparticles biodegrade after pro-survival drug delivery and ultrasound/magnetic resonance imaging of stem cells. Theranostics Kempen, P. J., Greasley, S., Parker, K. A., Campbell, J. L., Chang, H., Jones, J. R., Sinclair, R., Gambhir, S. S., Jokerst, J. V. 2015; 5 (6): 631-642

  Abstract

  Increasing cell survival in stem cell therapy is an important challenge for the field of regenerative medicine. Here, we report theranostic mesoporous silica nanoparticles that can increase cell survival through both diagnostic and therapeutic approaches. First, the nanoparticle offers ultrasound and MRI signal to guide implantation into the peri-infarct zone and away from the most necrotic tissue. Second, the nanoparticle serves as a slow release reservoir of insulin-like growth factor (IGF)-a protein shown to increase cell survival. Mesenchymal stem cells labeled with these nanoparticles had detection limits near 9000 cells with no cytotoxicity at the 250 µg/mL concentration required for labeling. We also studied the degradation of the nanoparticles and showed that they clear from cells in approximately 3 weeks. The presence of IGF increased cell survival up to 40% (p<0.05) versus unlabeled cells under in vitro serum-free culture conditions.

  View details for DOI 10.7150/thno.11389

  View details for PubMedID 25825602

• A correlative optical microscopy and scanning electron microscopy approach to locating nanoparticles in brain tumors. Micron Kempen, P. J., Kircher, M. F., de la Zerda, A., Zavaleta, C. L., Jokerst, J. V., Mellinghoff, I. K., Gambhir, S. S., Sinclair, R. 2015; 68: 70-76

  Abstract

  The growing use of nanoparticles in biomedical applications, including cancer diagnosis and treatment, demands the capability to exactly locate them within complex biological systems. In this work a correlative optical and scanning electron microscopy technique was developed to locate and observe multi-modal gold core nanoparticle accumulation in brain tumor models. Entire brain sections from mice containing orthotopic brain tumors injected intravenously with nanoparticles were imaged using both optical microscopy to identify the brain tumor, and scanning electron microscopy to identify the individual nanoparticles. Gold-based nanoparticles were readily identified in the scanning electron microscope using backscattered electron imaging as bright spots against a darker background. This information was then correlated to determine the exact location of the nanoparticles within the brain tissue. The nanoparticles were located only in areas that contained tumor cells, and not in the surrounding healthy brain tissue. This correlative technique provides a powerful method to relate the macro- and micro-scale features visible in light microscopy with the nanoscale features resolvable in scanning electron microscopy.

  View details for DOI 10.1016/j.micron.2014.09.004

  View details for PubMedID 25464144

  View details for PubMedCentralID PMC4262686

• A multimodal imaging agent for intrinsic surface enhanced Raman scattering of biological tissue Conference on Plasmonics in Biology and Medicine XII Pohling, C. B., Campbell, J. L., Larson, T. A., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2015

  View details for DOI 10.1117/12.2077890

  View details for Web of Science ID 000353615600006

• Validation of 64Cu-DOTA-rituximab injection preparation under good manufacturing practices: a PET tracer for imaging of B-cell non-Hodgkin lymphoma. Molecular imaging Natarajan, A., Arksey, N., Iagaru, A., Chin, F. T., Gambhir, S. S. 2015; 14

  Abstract

  AbstractManufacturing of 64Cu-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA)-rituximab injection under good manufacturing practices (GMP) was validated for imaging of patients with CD20+ B-cell non-Hodgkin lymphoma. Rituximab was purified by size exclusion high performance liquid chromatography (HPLC) and conjugated to DOTA-mono-(N-hydroxysuccinimidyl) ester. 64CuCl2, buffers, reagents, and other raw materials were obtained as high-grade quality. Following a semi-automated synthesis of 64Cu-DOTA-rituximab, a series of quality control tests was performed. The product was further tested in vivo using micro-positron emission tomography/computed tomography (PET/CT) to assess targeting ability towards human CD20 in transgenic mice. Three batches of 64Cu-DOTA-rituximab final product were prepared as per GMP specifications. The radiolabeling yield from these batches was 93.1 ± 5.8%; these provided final product with radiopharmaceutical yield, purity, and specific activity of 59.2 ± 5.1% (0.9 ± 0.1 GBq of 64Cu), > 95% (by HPLC and radio-thin layer chromatography), and 229.4 ± 43.3 GBq/µmol (or 1.5 ± 0.3 MBq/µg), respectively. The doses passed apyrogenicity and human serum stability specifications, were sterile up to 14 days, and retained > 60% immunoreactivity. In vivo micro-PET/CT mouse images at 24 hours postinjection showed that the tracer targeted the intended sites of human CD20 expression. Thus, we have validated the manufacturing of GMP grade 64Cu-DOTA-rituximab for injection in the clinical setting.

  View details for DOI 10.2310/7290.2014.00055

  View details for PubMedID 25762106

• Syntheses and Discovery of a Novel Class of Cinnamic Hydroxamates as Histone Deacetylase Inhibitors by Multimodality Molecular Imaging in Living Subjects CANCER RESEARCH CHAN, C. T., Qi, J., Smith, W., Paranol, R., Mazitschek, R., West, N., Reeves, R., Chiosis, G., Schreiber, S. L., Bradner, J. E., Paulmurugan, R., Gambhir, S. S. 2014; 74 (24): 7475-7486

  Abstract

  Histone deacetylases (HDAC) that regulate gene expression are being explored as cancer therapeutic targets. In this study, we focused on HDAC6 based on its ability to inhibit cancerous Hsp90 chaperone activities by disrupting Hsp90/p23 interactions. To identify novel HDAC6 inhibitors, we used a dual-luciferase reporter system in cell culture and living mice by bioluminescence imaging (BLI). On the basis of existing knowledge, a library of hydrazone compounds was generated for screening by coupling cinnamic hydroxamates with aldehydes and ketones. Potency and selectivity were determined by in vitro HDAC profiling assays, with further evaluation to inhibit Hsp90(α/β)/p23 interactions by BLI. In this manner, we identified compound 1A12 as a dose-dependent inhibitor of Hsp90(α/β)/p23 interactions, UKE-1 myeloid cell proliferation, p21(waf1) upregulation, and acetylated histone H3 levels. 1A12 was efficacious in tumor xenografts expressing Hsp90(α)/p23 reporters relative to carrier control-treated mice as determined by BLI. Small animal (18)F-FDG PET/CT imaging on the same cohort showed that 1A12 also inhibited glucose metabolism relative to control subjects. Ex vivo analyses of tumor lysates showed that 1A12 administration upregulated acetylated-H3 by approximately 3.5-fold. Taken together, our results describe the discovery and initial preclinical validation of a novel selective HDAC inhibitor.

  View details for DOI 10.1158/0008-5472.CAN-14-0197

  View details for Web of Science ID 000346363900031

  View details for PubMedID 25320008

• Noninvasive Reporter Gene Imaging of Human Oct4 (Pluripotency) Dynamics During the Differentiation of Embryonic Stem Cells in Living Subjects MOLECULAR IMAGING AND BIOLOGY Ahn, B., Parashurama, N., Patel, M., Ziv, K., Bhaumik, S., Yaghoubi, S. S., Paulmurugan, R., Gambhir, S. S. 2014; 16 (6): 865-876

  Abstract

  Human pluripotency gene networks (PGNs), controlled in part by Oct4, are central to understanding pluripotent stem cells, but current fluorescent reporter genes (RGs) preclude noninvasive assessment of Oct4 dynamics in living subjects.To assess Oc4 activity noninvasively, we engineered a mouse embryonic stem cell line which encoded both a pOct4-hrluc (humanized renilla luciferase) reporter and a pUbi-hfluc2-gfp (humanized firefly luciferase 2 fused to green fluorescent protein) reporter.In cell culture, pOct4-hRLUC activity demonstrated a peak at 48 h (day 2) and significant downregulation by 72 h (day 3) (p=0.0001). Studies in living subjects demonstrated significant downregulation in pOct4-hRLUC activity between 12 and 144 h (p = 0.001) and between 12 and 168 h (p = 0.0003). pOct4-hRLUC signal dynamics after implantation was complex, characterized by transient upregulation after initial downregulation in all experiments (n = 10, p = 0.01). As expected, cell culture differentiation of the engineered mouse embryonic stem cell line demonstrated activation of mesendodermal, mesodermal, endodermal, and ectodermal master regulators of differentiation, indicating potency to form all three germ layers.We conclude that the Oct4-hrluc RG system enables noninvasive Oct4 imaging in cell culture and in living subjects.

  View details for DOI 10.1007/s11307-014-0744-1

  View details for Web of Science ID 000345281600014

• Noninvasive reporter gene imaging of human Oct4 (pluripotency) dynamics during the differentiation of embryonic stem cells in living subjects. Molecular imaging and biology Ahn, B., Parashurama, N., Patel, M., Ziv, K., Bhaumik, S., Yaghoubi, S. S., Paulmurugan, R., Gambhir, S. S. 2014; 16 (6): 865-876

  Abstract

  Human pluripotency gene networks (PGNs), controlled in part by Oct4, are central to understanding pluripotent stem cells, but current fluorescent reporter genes (RGs) preclude noninvasive assessment of Oct4 dynamics in living subjects.To assess Oc4 activity noninvasively, we engineered a mouse embryonic stem cell line which encoded both a pOct4-hrluc (humanized renilla luciferase) reporter and a pUbi-hfluc2-gfp (humanized firefly luciferase 2 fused to green fluorescent protein) reporter.In cell culture, pOct4-hRLUC activity demonstrated a peak at 48 h (day 2) and significant downregulation by 72 h (day 3) (p=0.0001). Studies in living subjects demonstrated significant downregulation in pOct4-hRLUC activity between 12 and 144 h (p = 0.001) and between 12 and 168 h (p = 0.0003). pOct4-hRLUC signal dynamics after implantation was complex, characterized by transient upregulation after initial downregulation in all experiments (n = 10, p = 0.01). As expected, cell culture differentiation of the engineered mouse embryonic stem cell line demonstrated activation of mesendodermal, mesodermal, endodermal, and ectodermal master regulators of differentiation, indicating potency to form all three germ layers.We conclude that the Oct4-hrluc RG system enables noninvasive Oct4 imaging in cell culture and in living subjects.

  View details for DOI 10.1007/s11307-014-0744-1

  View details for PubMedID 24845530

• (18)F-FPPRGD2 PET/CT: pilot phase evaluation of breast cancer patients. Radiology Iagaru, A., Mosci, C., Shen, B., Chin, F. T., Mittra, E., Telli, M. L., Gambhir, S. S. 2014; 273 (2): 549-559

  Abstract

  Purpose To present data from the first prospective pilot phase trial of breast cancer participants imaged with fluorine 18 ((18)F)-2-fluoropropionyl-labeled PEGylated dimeric arginine-glycine-aspartic acid (RGD) peptide (PEG3-E[c{RGDyk}]2) (FPPRGD2), a radiopharmaceutical agent used in positron emission tomographic (PET) imaging. Materials and Methods The local institutional review board approved the HIPAA-compliant protocol. Written informed consent was obtained from each patient. Eight women (age range, 44-67 years; mean age, 54.3 years ± 8.8 [standard deviation]) with newly diagnosed or recurrent breast cancer were recruited between November 2010 and February 2011. (18)F-FPPRGD2 PET/computed tomographic (CT) and (18)F-fluorodeoxyglucose (FDG) PET/CT examinations were performed within 3 weeks of each other. Dynamic (18)F-FPPRGD2 PET and two whole-body static (18)F-FPPRGD2 PET/CT scans were obtained. During this time, vital signs and electrocardiograms were recorded at regular intervals. Blood samples were obtained before the injection of (18)F-FPPRGD2 and at 24 hours and 1 week after injection to evaluate for toxicity. A nonparametric version of multivariate analysis of variance was used to assess the safety outcome measures simultaneously across time points. A paired two-sample t test was performed to compare the maximum standardized uptake values (SUVmax). Results (18)F-FPPRGD2 was well tolerated, without noticeable changes in vital signs, on electrocardiograms, or in laboratory values. A total of 30 lesions were evaluated at (18)F-FDG PET/CT and (18)F-FPPRGD2 PET/CT. The primary breast lesions had (18)F-FPPRGD2 uptake with SUVmax of 2.4-9.4 (mean, 5.6 ± 2.8) 60 minutes after injection, compared with (18)F-FDG uptake with SUVmax of 2.8-18.6 (mean, 10.4 ± 7.2). Metastatic lesions also showed (18)F-FPPRGD2 uptake, with SUVmax of 2.4-9.7 (mean, 5.0 ± 2.3) at 60 minutes, compared with (18)F-FDG uptake with SUVmax of 2.2-14.6 (mean, 6.6 ± 4.2). Conclusion Data from this pilot phase study suggest that (18)F-FPPRGD2 is a safe PET radiopharmaceutical agent. Evaluation of (18)F-FPPRGD2 in participants with breast cancer demonstrated significant uptake in the primary lesion and in the metastases. Larger cohorts are required to confirm these preliminary findings. © RSNA, 2014.

  View details for DOI 10.1148/radiol.14140028

  View details for PubMedID 25033190

• THE NATURALLY OCCURRING STEROID, WITHAFERIN A, IN SYNERGISTIC CONCERT WITH HER2/EGFR INHIBITORS ABROGATES PROLIFERATION OF HUMAN GLIOBLASTOMA CELL CULTURES AT NANOMOLAR CONCENTRATIONS Chang, E., Abbasi, T., D'Souza, A., Gowrishankar, G., Mallick, P., Gambhir, S. S. OXFORD UNIV PRESS INC. 2014

  View details for DOI 10.1093/neuonc/nou246.3

  View details for Web of Science ID 000350452200246

• Cerenkov luminescence endoscopy: improved molecular sensitivity with ß--emitting radiotracers. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Carpenter, C. M., Ma, X., Liu, H., Sun, C., Pratx, G., Wang, J., Gambhir, S. S., Xing, L., Cheng, Z. 2014; 55 (11): 1905-1909

  Abstract

  Cerenkov luminescence endoscopy (CLE) is an optical technique that captures the Cerenkov photons emitted from highly energetic moving charged particles (β(+) or β(-)) and can be used to monitor the distribution of many clinically available radioactive probes. A main limitation of CLE is its limited sensitivity to small concentrations of radiotracer, especially when used with a light guide. We investigated the improvement in the sensitivity of CLE brought about by using a β(-) radiotracer that improved Cerenkov signal due to both higher β-particle energy and lower γ noise in the imaging optics because of the lack of positron annihilation.The signal-to-noise ratio (SNR) of (90)Y was compared with that of (18)F in both phantoms and small-animal tumor models. Sensitivity and noise characteristics were demonstrated using vials of activity both at the surface and beneath 1 cm of tissue. Rodent U87MG glioma xenograft models were imaged with radiotracers bound to arginine-glycine-aspartate (RGD) peptides to determine the SNR.γ noise from (18)F was demonstrated by both an observed blurring across the field of view and a more pronounced fall-off with distance. A decreased γ background and increased energy of the β particles resulted in a 207-fold improvement in the sensitivity of (90)Y compared with (18)F in phantoms. (90)Y-bound RGD peptide produced a higher tumor-to-background SNR than (18)F in a mouse model.The use of (90)Y for Cerenkov endoscopic imaging enabled superior results compared with an (18)F radiotracer.

  View details for DOI 10.2967/jnumed.114.139105

  View details for PubMedID 25300598

• Endometrial VEGF induces placental sFLT1 and leads to pregnancy complications JOURNAL OF CLINICAL INVESTIGATION Fan, X., Rai, A., Kambham, N., Sung, J. F., Singh, N., Petitt, M., Dhal, S., Agrawal, R., Sutton, R. E., Druzin, M. L., Gambhir, S. S., Ambati, B. K., Cross, J. C., Nayak, N. R. 2014; 124 (11): 4941-4952

  Abstract

  There is strong evidence that overproduction of soluble fms-like tyrosine kinase-1 (sFLT1) in the placenta is a major cause of vascular dysfunction in preeclampsia through sFLT1-dependent antagonism of VEGF. However, the cause of placental sFLT1 upregulation is not known. Here we demonstrated that in women with preeclampsia, sFLT1 is upregulated in placental trophoblasts, while VEGF is upregulated in adjacent maternal decidual cells. In response to VEGF, expression of sFlt1 mRNA, but not full-length Flt1 mRNA, increased in cultured murine trophoblast stem cells. We developed a method for transgene expression specifically in mouse endometrium and found that endometrial-specific VEGF overexpression induced placental sFLT1 production and elevated sFLT1 levels in maternal serum. This led to pregnancy losses, placental vascular defects, and preeclampsia-like symptoms, including hypertension, proteinuria, and glomerular endotheliosis in the mother. Knockdown of placental sFlt1 with a trophoblast-specific transgene caused placental vascular changes that were consistent with excess VEGF activity. Moreover, sFlt1 knockdown in VEGF-overexpressing animals enhanced symptoms produced by VEGF overexpression alone. These findings indicate that sFLT1 plays an essential role in maintaining vascular integrity in the placenta by sequestering excess maternal VEGF and suggest that a local increase in VEGF can trigger placental overexpression of sFLT1, potentially contributing to the development of preeclampsia and other pregnancy complications.

  View details for DOI 10.1172/JCI76864

  View details for Web of Science ID 000344203300029

  View details for PubMedCentralID PMC4347223

• F-18-FPPRGD2 PET/CT: Pilot Phase Evaluation of Breast Cancer Patients RADIOLOGY Lagaru, A., Mosci, C., Shen, B., Chin, F. T., Mittra, E., Telli, M. L., Gambhir, S. S. 2014; 273 (2): 549-559

  Abstract

  Purpose To present data from the first prospective pilot phase trial of breast cancer participants imaged with fluorine 18 ((18)F)-2-fluoropropionyl-labeled PEGylated dimeric arginine-glycine-aspartic acid (RGD) peptide (PEG3-E[c{RGDyk}]2) (FPPRGD2), a radiopharmaceutical agent used in positron emission tomographic (PET) imaging. Materials and Methods The local institutional review board approved the HIPAA-compliant protocol. Written informed consent was obtained from each patient. Eight women (age range, 44-67 years; mean age, 54.3 years ± 8.8 [standard deviation]) with newly diagnosed or recurrent breast cancer were recruited between November 2010 and February 2011. (18)F-FPPRGD2 PET/computed tomographic (CT) and (18)F-fluorodeoxyglucose (FDG) PET/CT examinations were performed within 3 weeks of each other. Dynamic (18)F-FPPRGD2 PET and two whole-body static (18)F-FPPRGD2 PET/CT scans were obtained. During this time, vital signs and electrocardiograms were recorded at regular intervals. Blood samples were obtained before the injection of (18)F-FPPRGD2 and at 24 hours and 1 week after injection to evaluate for toxicity. A nonparametric version of multivariate analysis of variance was used to assess the safety outcome measures simultaneously across time points. A paired two-sample t test was performed to compare the maximum standardized uptake values (SUVmax). Results (18)F-FPPRGD2 was well tolerated, without noticeable changes in vital signs, on electrocardiograms, or in laboratory values. A total of 30 lesions were evaluated at (18)F-FDG PET/CT and (18)F-FPPRGD2 PET/CT. The primary breast lesions had (18)F-FPPRGD2 uptake with SUVmax of 2.4-9.4 (mean, 5.6 ± 2.8) 60 minutes after injection, compared with (18)F-FDG uptake with SUVmax of 2.8-18.6 (mean, 10.4 ± 7.2). Metastatic lesions also showed (18)F-FPPRGD2 uptake, with SUVmax of 2.4-9.7 (mean, 5.0 ± 2.3) at 60 minutes, compared with (18)F-FDG uptake with SUVmax of 2.2-14.6 (mean, 6.6 ± 4.2). Conclusion Data from this pilot phase study suggest that (18)F-FPPRGD2 is a safe PET radiopharmaceutical agent. Evaluation of (18)F-FPPRGD2 in participants with breast cancer demonstrated significant uptake in the primary lesion and in the metastases. Larger cohorts are required to confirm these preliminary findings. © RSNA, 2014.

  View details for DOI 10.1148/radiol.14140028

  View details for Web of Science ID 000345069800028

• A Radiofluorinated Divalent Cystine Knot Peptide for Tumor PET Imaging MOLECULAR PHARMACEUTICS Jiang, L., Kimura, R. H., Ma, X., Tu, Y., Miao, Z., Shen, B., Chin, F. T., Shi, H., Gambhir, S. S., Cheng, Z. 2014; 11 (11): 3885-3892

  Abstract

  A divalent knottin containing two separate integrin binding epitopes (RGD) in the adjacent loops, 3-4A, was recently developed and reported in our previous publication. In the current study, 3-4A was radiofluorinated with a 4-nitrophenyl 2-(18)F-fluoropropinate ((18)F-NFP) group and the resulting divalent positron emission tomography (PET) probe, (18)F-FP-3-4A, was evaluated as a novel imaging probe to detect integrin αvβ3 positive tumors in living animals. Knottin 3-4A was synthesized by solid phase peptide synthesis, folded, and site-specifically conjugated with (18/19)F-NFP to produce the fluorinated peptide (18/19)F-fluoropropinate-3-4A ((18/19)F-FP-3-4A). The stability of (18)F-FP-3-4A was tested in both phosphate buffered saline (PBS) buffer and mouse serum. Cell uptake assays of the radiolabeled peptides were performed using U87MG cells. In addition, small animal PET imaging and biodistribution studies of (18)F-FP-3-4A were performed in U87MG tumor-bearing mice. The receptor targeting specificity of the radiolabeled peptide was also verified by coinjecting the probe with a blocking peptide cyclo(RGDyK). Our study showed that (18)F-FP-3-4A exhibited excellent stability in PBS buffer (pH 7.4) and mouse serum. Small animal PET imaging and biodistribution data revealed that (18)F-FP-3-4A exhibited rapid and good tumor uptake (3.76 ± 0.59% ID/g and 2.22 ± 0.62% ID/g at 0.5 and 1 h, respectively). (18)F-FP-3-4A was rapidly cleared from the normal tissues, resulting in excellent tumor-to-normal tissue contrasts. For example, liver uptake was only 0.39 ± 0.07% ID/g and the tumor to liver ratio was 5.69 at 1 h p.i. Furthermore, coinjection of cyclo(RGDyK) with (18)F-FP-3-4A significantly inhibited tumor uptake (0.41 ± 0.12 vs 1.02 ± 0.19% ID/g at 2.5 h) in U87MG xenograft models, demonstrating specific accumulation of the probe in the tumor. In summary, the divalent probe (18)F-FP-3-4A is characterized by rapid and high tumor uptake and excellent tumor-to-normal tissue ratios. (18)F-FP-3-4A is a highly promising knottin based PET probe for translating into clinical imaging of tumor angiogenesis.

  View details for DOI 10.1021/mp500018s

  View details for Web of Science ID 000344307700012

• Endoscopic molecular imaging of human bladder cancer using a CD47 antibody SCIENCE TRANSLATIONAL MEDICINE Pan, Y., Volkmer, J., Mach, K. E., Rouse, R. V., Liu, J., Sahoo, D., Chang, T. C., Metzner, T. J., Kang, L., van de Rijn, M., Skinner, E. C., Gambhir, S. S., Weissman, I. L., Liao, J. C. 2014; 6 (260)

  Abstract

  A combination of optical imaging technologies with cancer-specific molecular imaging agents is a potentially powerful strategy to improve cancer detection and enable image-guided surgery. Bladder cancer is primarily managed endoscopically by white light cystoscopy with suboptimal diagnostic accuracy. Emerging optical imaging technologies hold great potential for improved diagnostic accuracy but lack imaging agents for molecular specificity. Using fluorescently labeled CD47 antibody (anti-CD47) as molecular imaging agent, we demonstrated consistent identification of bladder cancer with clinical grade fluorescence imaging systems, confocal endomicroscopy, and blue light cystoscopy in fresh surgically removed human bladders. With blue light cystoscopy, the sensitivity and specificity for CD47-targeted imaging were 82.9 and 90.5%, respectively. We detected variants of bladder cancers, which are diagnostic challenges, including carcinoma in situ, residual carcinoma in tumor resection bed, recurrent carcinoma following prior intravesical immunotherapy with Bacillus Calmette-Guérin (BCG), and excluded cancer from benign but suspicious-appearing mucosa. CD47-targeted molecular imaging could improve diagnosis and resection thoroughness for bladder cancer.

  View details for DOI 10.1126/scitranslmed.3009457

  View details for Web of Science ID 000343920500006

• Endoscopic molecular imaging of human bladder cancer using a CD47 antibody. Science translational medicine Pan, Y., Volkmer, J., Mach, K. E., Rouse, R. V., Liu, J., Sahoo, D., Chang, T. C., Metzner, T. J., Kang, L., van de Rijn, M., Skinner, E. C., Gambhir, S. S., Weissman, I. L., Liao, J. C. 2014; 6 (260): 260ra148-?

  Abstract

  A combination of optical imaging technologies with cancer-specific molecular imaging agents is a potentially powerful strategy to improve cancer detection and enable image-guided surgery. Bladder cancer is primarily managed endoscopically by white light cystoscopy with suboptimal diagnostic accuracy. Emerging optical imaging technologies hold great potential for improved diagnostic accuracy but lack imaging agents for molecular specificity. Using fluorescently labeled CD47 antibody (anti-CD47) as molecular imaging agent, we demonstrated consistent identification of bladder cancer with clinical grade fluorescence imaging systems, confocal endomicroscopy, and blue light cystoscopy in fresh surgically removed human bladders. With blue light cystoscopy, the sensitivity and specificity for CD47-targeted imaging were 82.9 and 90.5%, respectively. We detected variants of bladder cancers, which are diagnostic challenges, including carcinoma in situ, residual carcinoma in tumor resection bed, recurrent carcinoma following prior intravesical immunotherapy with Bacillus Calmette-Guérin (BCG), and excluded cancer from benign but suspicious-appearing mucosa. CD47-targeted molecular imaging could improve diagnosis and resection thoroughness for bladder cancer.

  View details for DOI 10.1126/scitranslmed.3009457

  View details for PubMedID 25355698

• Transferring biomarker into molecular probe: melanin nanoparticle as a naturally active platform for multimodality imaging. Journal of the American Chemical Society Fan, Q., Cheng, K., Hu, X., Ma, X., Zhang, R., Yang, M., Lu, X., Xing, L., Huang, W., Gambhir, S. S., Cheng, Z. 2014; 136 (43): 15185-15194

  Abstract

  Developing multifunctional and easily prepared nanoplatforms with integrated different modalities is highly challenging for molecular imaging. Here, we report the successful transferring an important molecular target, melanin, into a novel mul-timodality imaging nanoplatform. Melanin is abundantly expressed in melanotic melanomas and thus has been actively studied as a target for melanoma imaging. In our work, the multifunctional biopolymer nanoplatform based on ultrasmall (< 10 nm) water-soluble melanin nanoparticle (MNP) was developed and showed unique photoacoustic property and natural binding ability with metal ions (for example, 64Cu2+, Fe3+). Therefore MNP can serve not only as a photoacoustic contrast agent, but also as a nanoplatform for positron emission tomography (PET) and magnetic resonance imaging (MRI). Traditional passive nanoplatforms require complicated and time-consuming processes for pre-building reporting moieties or chemical modifications using active groups to integrate different contrast properties into one entity. In comparison, utilizing functional biomarker melanin can greatly simplify the building process. We further conjugated αvβ3 integrins targeting peptide, cyclic c(RGDfC) peptide, to MNPs and this allowed targeting of these nanoparticles to allow for greater U87MG tumor accumulation than that simply possible due to the enhanced permeability and retention (EPR) effect. The multimodal properties of MNPs demonstrate the high potential of endogenous materials with multifunctions as nanoplatforms for molecular theranostics and clinical translation.

  View details for DOI 10.1021/ja505412p

  View details for PubMedID 25292385

• Correlation of plasma biomarker levels with early-stage tumor viability in an orthotopic ovarian cancer mouse model Hori, S. S., Lutz, A. M., Paulmurugan, R., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2014

  View details for DOI 10.1158/1538-7445.AM2014-873

  View details for Web of Science ID 000349906905331

• MicroRNA footprints of circulating tumor cells in patients with non-small cell lung cancer Nair, V. S., Giraldez, M., Luttgen, M., Keu, K., Vasanawala, M., Horng, G., Jamali, M., Kolatkar, A., Kuschner, W., Kuhn, P., Gambhir, S., Tewari, M. AMER ASSOC CANCER RESEARCH. 2014

  View details for DOI 10.1158/1538-7445.AM2014-4006

  View details for Web of Science ID 000349910201490

• Novel method of liver tumor detection and characterization using ultrasound-induced biomarker release D'Souza, A. L., Yan, X., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2014

  View details for DOI 10.1158/1538-7445.AM2014-4287

  View details for Web of Science ID 000349910202263

• First evaluation of a time-of-flight whole-body PET/MRI scanner in oncology patients: comparison with PET/CT Iagaru, A., Jamali, M., Minamimoto, R., Mittra, E., Gold, G., Vasanwala, S., Gambhir, S. S., Zaharchuk, G. SPRINGER. 2014: S287–S288

  View details for Web of Science ID 000348841900416

• Molecular photoacoustic imaging and serum diagnostics rapidly detect response to angiopoietin 1 and 2 blockade in ovarian cancer Bohndiek, S. E., Sasportas, L., Machtaler, S., Jokerst, J. V., Hori, S., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2014

  View details for DOI 10.1158/1538-7445.AM2014-2047

  View details for Web of Science ID 000349906902259

• Investigation of 6-[F-18]-Fluoromaltose as a Novel PET Tracer for Imaging Bacterial Infection PLOS ONE Gowrishankar, G., Namavari, M., Jouannot, E. B., Hoehne, A., Reeves, R., Hardy, J., Gambhir, S. S. 2014; 9 (9)

  Abstract

  Despite advances in the field of nuclear medicine, the imaging of bacterial infections has remained a challenge. The existing reagents suffer from poor sensitivity and specificity. In this study we investigate the potential of a novel PET (positron emission tomography) tracer that overcomes these limitations.6-[¹⁸F]-fluoromaltose was synthesized. Its behavior in vitro was evaluated in bacterial and mammalian cultures. Detailed pharmacokinetic and biodistribution profiles for the tracer were obtained from a murine model.6-[¹⁸F]-fluoromaltose is taken up by multiple strains of pathogenic bacteria. It is not taken up by mammalian cancer cell lines. 6-[¹⁸F]-fluoromaltose is retained in infected muscles in a murine model of bacterial myositis. It does not accumulate in inflamed tissue.We have shown that 6-[¹⁸F]-fluoromaltose can be used to image bacterial infection in vivo with high specificity. We believe that this class of agents will have a significant impact on the clinical management of patients.

  View details for DOI 10.1371/journal.pone.0107951

  View details for Web of Science ID 000343679800058

  View details for PubMedCentralID PMC4171493

• Cellulose Nanoparticles are a Biodegradable Photoacoustic Contrast Agent for Use in Living Mice. Photoacoustics Jokerst, J. V., Van de Sompel, D., Bohndiek, S. E., Gambhir, S. S. 2014; 2 (3): 119-127

  Abstract

  Molecular imaging with photoacoustic ultrasound is an emerging field that combines the spatial and temporal resolution of ultrasound with the contrast of optical imaging. However, there are few imaging agents that offer both high signal intensity and biodegradation into small molecules. Here we describe a cellulose-based nanoparticle with peak photoacoustic signal at 700 nm and an in vitro limit of detection of 6 pM (0.02 mg/mL). Doses down to 0.35 nM (1.2 mg/mL) were used to image mouse models of ovarian cancer. Most importantly, the nanoparticles were shown to biodegrade in the presence of cellulase both through a glucose assay and electron microscopy.

  View details for PubMedID 25225633

• Cellulose nanoparticles are a biodegradable photoacoustic contrast agent for use in living mice PHOTOACOUSTICS Jokerst, J. V., Van de Sompel, D., Bohndiek, S. E., Gambhir, S. S. 2014; 2 (3): 119–27

  View details for DOI 10.1016/j.pacs.2014.07.001

  View details for Web of Science ID 000218887500003

• Circulating Tumor Microemboli Diagnostics for Patients with Non-Small-Cell Lung Cancer JOURNAL OF THORACIC ONCOLOGY Carlsson, A., Nair, V. S., Luttgen, M. S., Keu, K. V., Horng, G., Vasanawala, M., Kolatkar, A., Jamali, M., Iagaru, A. H., Kuschner, W., Loo, B. W., Shrager, J. B., Bethel, K., Hoh, C. K., Bazhenova, L., Nieva, J., Kuhn, P., Gambhir, S. S. 2014; 9 (8): 1111-1119

  Abstract

  Circulating tumor microemboli (CTM) are potentially important cancer biomarkers, but using them for cancer detection in early-stage disease has been assay limited. We examined CTM test performance using a sensitive detection platform to identify stage I non-small-cell lung cancer (NSCLC) patients undergoing imaging evaluation.First, we prospectively enrolled patients during 18F-FDG PET-CT imaging evaluation for lung cancer that underwent routine phlebotomy where CTM and circulating tumor cells (CTCs) were identified in blood using nuclear (DAPI), cytokeratin (CK), and CD45 immune-fluorescent antibodies followed by morphologic identification. Second, CTM and CTC data were integrated with patient (age, gender, smoking, and cancer history) and imaging (tumor diameter, location in lung, and maximum standard uptake value [SUVmax]) data to develop and test multiple logistic regression models using a case-control design in a training and test cohort followed by cross-validation in the entire group.We examined 104 patients with NSCLC, and the subgroup of 80 with stage I disease, and compared them to 25 patients with benign disease. Clinical and imaging data alone were moderately discriminating for all comers (Area under the Curve [AUC] = 0.77) and by stage I disease only (AUC = 0.77). However, the presence of CTM combined with clinical and imaging data was significantly discriminating for diagnostic accuracy in all NSCLC patients (AUC = 0.88, p value = 0.001) and for stage I patients alone (AUC = 0.87, p value = 0.002).CTM may add utility for lung cancer diagnosis during imaging evaluation using a sensitive detection platform.

  View details for PubMedID 25157764

• Imaging of hepatocellular carcinoma patient-derived xenografts using (89)Zr-labeled anti-glypican-3 monoclonal antibody. Biomaterials Yang, X., Liu, H., Sun, C. K., Natarajan, A., Hu, X., Wang, X., Allegretta, M., Guttmann, R. D., Gambhir, S. S., Chua, M. S., Cheng, Z., So, S. K. 2014; 35 (25): 6964-71

  Abstract

  Imaging probes for early detection of hepatocellular carcinoma (HCC) are highly desired to overcome current diagnostic limitations which lead to poor prognosis. The membrane protein glypican-3 (GPC3) is a potential molecular target for early HCC detection as it is over-expressed in >50% of HCCs, and is associated with early hepatocarcinogenesis. We synthesized the positron emission tomography (PET) probe (89)Zr-DFO-1G12 by bioconjugating and radiolabeling the anti-GPC3 monoclonal antibody (clone 1G12) with (89)Zr, and evaluated its tumor-targeting capacity. In vitro, (89)Zr-DFO-1G12 was specifically taken up into GPC3-positive HCC cells only, but not in the GPC3-negative prostate cancer cell line (PC3). In vivo, (89)Zr-DFO-1G12 specifically accumulated in subcutaneous GPC3-positive HCC xenografts only, but not in PC3 xenografts. Importantly, (89)Zr-DFO-1G12 delineated orthotopic HCC xenografts from surrounding normal liver, with tumor/liver (T/L) ratios of 6.65 ± 1.33 for HepG2, and 4.29 ± 0.52 for Hep3B xenografts. It also delineated orthotopic xenografts derived from three GPC3-positive HCC patient specimens, with T/L ratios of 4.21 ± 0.64, 2.78 ± 0.26, and 2.31 ± 0.38 at 168 h p.i. Thus, (89)Zr-DFO-1G12 is a highly translatable probe for the specific and high contrast imaging of GPC3-positive HCCs, which may aid early detection of HCC to allow timely intervention.

  View details for DOI 10.1016/j.biomaterials.2014.04.089

  View details for PubMedID 24836949

• Imaging of hepatocellular carcinoma patient-derived xenografts using Zr-89-labeled anti-glypican-3 monoclonal antibody BIOMATERIALS Yang, X., Liu, H., Sun, C. K., Natarajan, A., Hu, X., Wang, X., Allegretta, M., Guttmann, R. D., Gambhir, S. S., Chua, M., Cheng, Z., So, S. K. 2014; 35 (25): 6964-6971

  Abstract

  Imaging probes for early detection of hepatocellular carcinoma (HCC) are highly desired to overcome current diagnostic limitations which lead to poor prognosis. The membrane protein glypican-3 (GPC3) is a potential molecular target for early HCC detection as it is over-expressed in >50% of HCCs, and is associated with early hepatocarcinogenesis. We synthesized the positron emission tomography (PET) probe (89)Zr-DFO-1G12 by bioconjugating and radiolabeling the anti-GPC3 monoclonal antibody (clone 1G12) with (89)Zr, and evaluated its tumor-targeting capacity. In vitro, (89)Zr-DFO-1G12 was specifically taken up into GPC3-positive HCC cells only, but not in the GPC3-negative prostate cancer cell line (PC3). In vivo, (89)Zr-DFO-1G12 specifically accumulated in subcutaneous GPC3-positive HCC xenografts only, but not in PC3 xenografts. Importantly, (89)Zr-DFO-1G12 delineated orthotopic HCC xenografts from surrounding normal liver, with tumor/liver (T/L) ratios of 6.65 ± 1.33 for HepG2, and 4.29 ± 0.52 for Hep3B xenografts. It also delineated orthotopic xenografts derived from three GPC3-positive HCC patient specimens, with T/L ratios of 4.21 ± 0.64, 2.78 ± 0.26, and 2.31 ± 0.38 at 168 h p.i. Thus, (89)Zr-DFO-1G12 is a highly translatable probe for the specific and high contrast imaging of GPC3-positive HCCs, which may aid early detection of HCC to allow timely intervention.

  View details for DOI 10.1016/j.biomaterials.2014.04.089

  View details for Web of Science ID 000338386800028

• A High-Affinity, High-Stability Photoacoustic Agent for Imaging Gastrin-Releasing Peptide Receptor in Prostate Cancer CLINICAL CANCER RESEARCH Levi, J., Sathirachinda, A., Gambhir, S. S. 2014; 20 (14): 3721-3729

  Abstract

  To evaluate the utility of targeted photoacoustic imaging (PAI) in providing molecular information to complement intrinsic functional and anatomical details of the vasculature within prostate lesion.We developed a PAI agent, AA3G-740, that targets gastrin-releasing peptide receptor (GRPR), found to be highly overexpressed in prostate cancer. The binding specificity of the agent was evaluated in human prostate cancer cell lines, PC3 and LNCaP, and antagonist properties determined by cell internalization and intracellular calcium mobilization studies. The imaging sensitivity was assessed for the agent itself and for the PC3 cells labeled with agent. The in vivo stability of the agent was determined in human plasma and in the blood of living mice. The in vivo binding of the agent was evaluated in PC3 prostate tumor models in mice, and was validated ex vivo by optical imaging.AA3G-740 demonstrated strong and specific binding to GRPR. The sensitivity of detection in vitro indicated suitability of the agent to image very small lesions. In mice, the agent was able to bind to GRPR even in poorly vascularized tumors leading to nearly 2-fold difference in photoacoustic signal relative to the control agent.The ability to image both vasculature and molecular profile outside the blood vessels gives molecular PAI a unique advantage over currently used imaging techniques. The imaging method presented here can find application both in diagnosis and in image-guided biopsy.

  View details for DOI 10.1158/1078-0432.CCR-13-3405

  View details for Web of Science ID 000339611500013

  View details for PubMedID 24850845

  View details for PubMedCentralID PMC4121111

• Selective uptake of single-walled carbon nanotubes by circulating monocytes for enhanced tumour delivery. Nature nanotechnology Smith, B. R., Ghosn, E. E., Rallapalli, H., Prescher, J. A., Larson, T., Herzenberg, L. A., Gambhir, S. S. 2014; 9 (6): 481-487

  Abstract

  In cancer imaging, nanoparticle biodistribution is typically visualized in living subjects using 'bulk' imaging modalities such as magnetic resonance imaging, computerized tomography and whole-body fluorescence. Accordingly, nanoparticle influx is observed only macroscopically, and the mechanisms by which they target cancer remain elusive. Nanoparticles are assumed to accumulate via several targeting mechanisms, particularly extravasation (leakage into tumour). Here, we show that, in addition to conventional nanoparticle-uptake mechanisms, single-walled carbon nanotubes are almost exclusively taken up by a single immune cell subset, Ly-6C(hi) monocytes (almost 100% uptake in Ly-6C(hi) monocytes, below 3% in all other circulating cells), and delivered to the tumour in mice. We also demonstrate that a targeting ligand (RGD) conjugated to nanotubes significantly enhances the number of single-walled carbon nanotube-loaded monocytes reaching the tumour (P < 0.001, day 7 post-injection). The remarkable selectivity of this tumour-targeting mechanism demonstrates an advanced immune-based delivery strategy for enhancing specific tumour delivery with substantial penetration.

  View details for DOI 10.1038/nnano.2014.62

  View details for PubMedID 24727688

• Prospective evaluation of combined NaF/FDG PET/CT and whole-body MRI in patients with breast and prostate cancer Iagaru, A., Mosci, C., Jamali, M., Loening, A., Mittra, E., Gambhir, S., Vasanawala, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438102203

• 99mTc-MDP scintigraphy vs. 18F NaF PET/CT for detection of skeletal metastases Iagaru, A., Jackson, T., Sabbah, N., Guo, H., Quon, A., Mittra, E., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438103152

• 99mTc-MDP scintigraphy vs. 18F NaF PET/CT for detection of skeletal metastases Iagaru, A., Jackson, T., Sabbah, N., Guo, H., Quon, A., Mittra, E., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438104142

• Single cell metabolomics in circulating tumor cells Sasportas, L., Turkcan, S., Pratx, G., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438100009

• Observed standardized uptake values in normal tissues and malignant lesions on combined 18F-NaF/18F-FDG PET/CT Minamimoto, R., Mosci, C., Jamali, M., Mittra, E., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438102304

• Combined NaF/FDG PET/CT evaluation of prostate cancer patients Iagaru, A., Mosci, C., Keu, K., Mittra, E., Hancock, S., Pachynski, R., Srinivas, S., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438102320

• 18F FPPRGD2 PET/CT evaluation of patients with suspected recurrence of glioblastoma multiforme Iagaru, A., Mosci, C., Jamali, M., Minamimoto, R., Mittra, E., Shen, B., Chin, F., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438100032

• A novel radioluminescence microscope for imaging radiotracers at the single-cell level Pratx, G., Natarajan, A., Turkcan, S., Sasportas, L., Axente, M., Gambhir, S., Xing, L. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438100043

• FDG uptake in normal tissues and malignant lesions from the first whole-body time-of-flight PET/MRI scanner: Comparison with PET/CT Iagaru, A., Mittra, E., Zaharchuk, G., Frost, R., Elekes, A., Anderson, J., Bobb, C., Lahrman, J., Gold, G., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438101156

• Tumor imaging with a radiofluorinated divalent knottin Jiang, L., Kimura, R., Ma, X., Tu, Y., Miao, Z., Shen, B., Chin, F., Shi, H., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438101248

• Detection of tumor cell apoptosis by PET: Comparison of [F-18]C-SNAT, [Tc-99m]HYNIC-Annexin V and [F-18]ML-10 Witney, T., Hoehne, A., Ilovich, O., Namavari, M., Shen, B., Chin, F., Rao, J., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2014

  View details for Web of Science ID 000361438102043

• A Radiofluorinated Divalent Cystine Knot Peptide for Tumor PET Imaging. Molecular pharmaceutics Jiang, L., Kimura, R. H., Ma, X., Tu, Y., Miao, Z., Shen, B., Chin, F. T., Shi, H., Gambhir, S. S., Cheng, Z. 2014

  Abstract

  A divalent knottin containing two separate integrin binding epitopes (RGD) in the adjacent loops, 3-4A, was recently developed and reported in our previous publication. In the current study, 3-4A was radiofluorinated with a 4-nitrophenyl 2-(18)F-fluoropropinate ((18)F-NFP) group and the resulting divalent positron emission tomography (PET) probe, (18)F-FP-3-4A, was evaluated as a novel imaging probe to detect integrin αvβ3 positive tumors in living animals. Knottin 3-4A was synthesized by solid phase peptide synthesis, folded, and site-specifically conjugated with (18/19)F-NFP to produce the fluorinated peptide (18/19)F-fluoropropinate-3-4A ((18/19)F-FP-3-4A). The stability of (18)F-FP-3-4A was tested in both phosphate buffered saline (PBS) buffer and mouse serum. Cell uptake assays of the radiolabeled peptides were performed using U87MG cells. In addition, small animal PET imaging and biodistribution studies of (18)F-FP-3-4A were performed in U87MG tumor-bearing mice. The receptor targeting specificity of the radiolabeled peptide was also verified by coinjecting the probe with a blocking peptide cyclo(RGDyK). Our study showed that (18)F-FP-3-4A exhibited excellent stability in PBS buffer (pH 7.4) and mouse serum. Small animal PET imaging and biodistribution data revealed that (18)F-FP-3-4A exhibited rapid and good tumor uptake (3.76 ± 0.59% ID/g and 2.22 ± 0.62% ID/g at 0.5 and 1 h, respectively). (18)F-FP-3-4A was rapidly cleared from the normal tissues, resulting in excellent tumor-to-normal tissue contrasts. For example, liver uptake was only 0.39 ± 0.07% ID/g and the tumor to liver ratio was 5.69 at 1 h p.i. Furthermore, coinjection of cyclo(RGDyK) with (18)F-FP-3-4A significantly inhibited tumor uptake (0.41 ± 0.12 vs 1.02 ± 0.19% ID/g at 2.5 h) in U87MG xenograft models, demonstrating specific accumulation of the probe in the tumor. In summary, the divalent probe (18)F-FP-3-4A is characterized by rapid and high tumor uptake and excellent tumor-to-normal tissue ratios. (18)F-FP-3-4A is a highly promising knottin based PET probe for translating into clinical imaging of tumor angiogenesis.

  View details for DOI 10.1021/mp500018s

  View details for PubMedID 24717098

• 99mTc-labeled cystine knot peptide targeting integrin avß6 for tumor SPECT imaging. Molecular pharmaceutics Zhu, X., Li, J., Hong, Y., Kimura, R. H., Ma, X., Liu, H., Qin, C., Hu, X., Hayes, T. R., Benny, P., Gambhir, S. S., Cheng, Z. 2014; 11 (4): 1208-1217

  Abstract

  Integrin αvβ6 is overexpressed in a variety of cancers, and its expression is often associated with poor prognosis. Therefore, there is a need to develop affinity reagents for noninvasive imaging of integrin αvβ6 expression since it may provide early cancer diagnosis, more accurate prognosis, and better treatment planning. We recently engineered and validated highly stable cystine knot peptides that selectively bind integrin αvβ6 with no cross-reactivity to integrins αvβ5, α5β1, or αvβ3, also known to be overexpressed in many cancers. Here, we developed a single photon emission computed tomography (SPECT) probe for imaging integrin αvβ6 positive tumors. Cystine knot peptide, S02, was first conjugated with a single amino acid chelate (SAAC) and labeled with [(99m)Tc(H2O)3(CO)3](+). The resulting probe, (99m)Tc-SAAC-S02, was then evaluated by in vitro cell uptake studies using two αvβ6 positive cell lines (human lung adenocarcinoma cell line HCC4006 and pancreatic cancer cell line BxPC-3) and two αvβ6 negative cell lines (human lung adenocarcinoma cell line H838 and human embryonic kidney cell line 293T). Next, SPECT/CT and biodistribution studies were performed in nude mice bearing HCC4006 and H838 tumor xenografts to evaluate the in vivo performance of (99m)Tc-SAAC-S02. Significant differences in the uptake of (99m)Tc-SAAC-S02 were observed in αvβ6 positive vs negative cells (P < 0.05). Biodistribution and small animal SPECT/CT studies revealed that (99m)Tc-SAAC-S02 accumulated to moderate levels in antigen positive tumors (∼2% ID/g at 1 and 6 h postinjection, n = 3 or 4/group). Moreover, the probe demonstrated tumor-to-background tissue ratios of 6.81 ± 2.32 (tumor-to-muscle) and 1.63 ± 0.18 (tumor-to-blood) at 6 h postinjection in αvβ6 positive tumor xenografts. Co-incubation of the probe with excess amount of unlabeled S02 as a blocking agent demonstrated significantly reduced tumor uptake, which is consistent with specific binding to the target. Renal filtration was the main route of clearance. In conclusion, knottin peptides are excellent scaffolds for which to develop highly stable imaging probes for a variety of oncological targets. (99m)Tc-SAAC-S02 demonstrates promise for use as a SPECT agent to image integrin αvβ6 expression in living systems.

  View details for DOI 10.1021/mp400683q

  View details for PubMedID 24524409

• A tunable silk-alginate hydrogel scaffold for stem cell culture and transplantation. Biomaterials Ziv, K., Nuhn, H., Ben-Haim, Y., Sasportas, L. S., Kempen, P. J., Niedringhaus, T. P., Hrynyk, M., Sinclair, R., Barron, A. E., Gambhir, S. S. 2014; 35 (12): 3736-3743

  Abstract

  One of the major challenges in regenerative medicine is the ability to recreate the stem cell niche, which is defined by its signaling molecules, the creation of cytokine gradients, and the modulation of matrix stiffness. A wide range of scaffolds has been developed in order to recapitulate the stem cell niche, among them hydrogels. This paper reports the development of a new silk-alginate based hydrogel with a focus on stem cell culture. This biocomposite allows to fine tune its elasticity during cell culture, addressing the importance of mechanotransduction during stem cell differentiation. The silk-alginate scaffold promotes adherence of mouse embryonic stem cells and cell survival upon transplantation. In addition, it has tunable stiffness as function of the silk-alginate ratio and the concentration of crosslinker - a characteristic that is very hard to accomplish in current hydrogels. The hydrogel and the presented results represents key steps on the way of creating artificial stem cell niche, opening up new paths in regenerative medicine.

  View details for DOI 10.1016/j.biomaterials.2014.01.029

  View details for PubMedID 24484675

• Tc-99m-Labeled Cystine Knot Peptide Targeting Integrin alpha(v)beta(6) for Tumor SPECT Imaging MOLECULAR PHARMACEUTICS Zhu, X., Li, J., Hong, Y., Kimura, R. H., Ma, X., Liu, H., Qin, C., Hu, X., Hayes, T. R., Benny, P., Gambhir, S. S., Cheng, Z. 2014; 11 (4): 1208-1217

  Abstract

  Integrin αvβ6 is overexpressed in a variety of cancers, and its expression is often associated with poor prognosis. Therefore, there is a need to develop affinity reagents for noninvasive imaging of integrin αvβ6 expression since it may provide early cancer diagnosis, more accurate prognosis, and better treatment planning. We recently engineered and validated highly stable cystine knot peptides that selectively bind integrin αvβ6 with no cross-reactivity to integrins αvβ5, α5β1, or αvβ3, also known to be overexpressed in many cancers. Here, we developed a single photon emission computed tomography (SPECT) probe for imaging integrin αvβ6 positive tumors. Cystine knot peptide, S02, was first conjugated with a single amino acid chelate (SAAC) and labeled with [(99m)Tc(H2O)3(CO)3](+). The resulting probe, (99m)Tc-SAAC-S02, was then evaluated by in vitro cell uptake studies using two αvβ6 positive cell lines (human lung adenocarcinoma cell line HCC4006 and pancreatic cancer cell line BxPC-3) and two αvβ6 negative cell lines (human lung adenocarcinoma cell line H838 and human embryonic kidney cell line 293T). Next, SPECT/CT and biodistribution studies were performed in nude mice bearing HCC4006 and H838 tumor xenografts to evaluate the in vivo performance of (99m)Tc-SAAC-S02. Significant differences in the uptake of (99m)Tc-SAAC-S02 were observed in αvβ6 positive vs negative cells (P < 0.05). Biodistribution and small animal SPECT/CT studies revealed that (99m)Tc-SAAC-S02 accumulated to moderate levels in antigen positive tumors (∼2% ID/g at 1 and 6 h postinjection, n = 3 or 4/group). Moreover, the probe demonstrated tumor-to-background tissue ratios of 6.81 ± 2.32 (tumor-to-muscle) and 1.63 ± 0.18 (tumor-to-blood) at 6 h postinjection in αvβ6 positive tumor xenografts. Co-incubation of the probe with excess amount of unlabeled S02 as a blocking agent demonstrated significantly reduced tumor uptake, which is consistent with specific binding to the target. Renal filtration was the main route of clearance. In conclusion, knottin peptides are excellent scaffolds for which to develop highly stable imaging probes for a variety of oncological targets. (99m)Tc-SAAC-S02 demonstrates promise for use as a SPECT agent to image integrin αvβ6 expression in living systems.

  View details for DOI 10.1021/mp400683q

  View details for Web of Science ID 000334092700013

  View details for PubMedCentralID PMC3993876

• A titratable two-step transcriptional amplification strategy for targeted gene therapy based on ligand-induced intramolecular folding of a mutant human estrogen receptor. Molecular imaging and biology Chen, I. Y., Paulmurugan, R., Nielsen, C. H., Wang, D. S., Chow, V., Robbins, R. C., Gambhir, S. S. 2014; 16 (2): 224-234

  Abstract

  The efficacy and safety of cardiac gene therapy depend critically on the level and the distribution of therapeutic gene expression following vector administration. We aimed to develop a titratable two-step transcriptional amplification (tTSTA) vector strategy, which allows modulation of transcriptionally targeted gene expression in the myocardium.We constructed a tTSTA plasmid vector (pcTnT-tTSTA-fluc), which uses the cardiac troponin T (cTnT) promoter to drive the expression of the recombinant transcriptional activator GAL4-mER(LBD)-VP2, whose ability to transactivate the downstream firefly luciferase reporter gene (fluc) depends on the binding of its mutant estrogen receptor (ER(G521T)) ligand binding domain (LBD) to an ER ligand such as raloxifene. Mice underwent either intramyocardial or hydrodynamic tail vein (HTV) injection of pcTnT-tTSTA-fluc, followed by differential modulation of fluc expression with varying doses of intraperitoneal raloxifene prior to bioluminescence imaging to assess the kinetics of myocardial or hepatic fluc expression.Intramyocardial injection of pcTnT-tTSTA-fluc followed by titration with intraperitoneal raloxifene led to up to tenfold induction of myocardial fluc expression. HTV injection of pcTnT-tTSTA-fluc led to negligible long-term hepatic fluc expression, regardless of the raloxifene dose given.The tTSTA vector strategy can effectively modulate transgene expression in a tissue-specific manner. Further refinement of this strategy should help maximize the benefit-to-risk ratio of cardiac gene therapy.

  View details for DOI 10.1007/s11307-013-0673-4

  View details for PubMedID 23955099

  View details for PubMedCentralID PMC4154804

• Construction of hybrid nanostructures as multimodal contrast agents for cancer imaging Cheng, K., Jokerst, J. V., Liu, H., Kothapalli, R., Jiang, H., Yang, M., Su, X., Ming, S., Hu, X., Li, J., Liu, Y., Levi, J., Gambhir, S., Cheng, Z. AMER CHEMICAL SOC. 2014

  View details for Web of Science ID 000348455204466

• Construction and validation of nano gold tripods for molecular imaging of living subjects. Journal of the American Chemical Society Cheng, K., Kothapalli, S., Liu, H., Koh, A. L., Jokerst, J. V., Jiang, H., Yang, M., Li, J., Levi, J., Wu, J. C., Gambhir, S. S., Cheng, Z. 2014; 136 (9): 3560-3571

  Abstract

  Anisotropic colloidal hybrid nanoparticles exhibit superior optical and physical properties compared to their counterparts with regular architectures. We herein developed a controlled, stepwise strategy to build novel, anisotropic, branched, gold nanoarchitectures (Au-tripods) with predetermined composition and morphology for bioimaging. The resultant Au-tripods with size less than 20 nm showed great promise as contrast agents for in vivo photoacoustic imaging (PAI). We further identified Au-tripods with two possible configurations as high-absorbance nanomaterials from various gold multipods using a numerical simulation analysis. The PAI signals were linearly correlated with their concentrations after subcutaneous injection. The in vivo biodistribution of Au-tripods favorable for molecular imaging was confirmed using small animal positron emission tomography (PET). Intravenous administration of cyclic Arg-Gly-Asp-d-Phe-Cys (RGDfC) peptide conjugated Au-tripods (RGD-Au-tripods) to U87MG tumor-bearing mice showed PAI contrasts in tumors almost 3-fold higher than for the blocking group. PAI results correlated well with the corresponding PET images. Quantitative biodistribution data revealed that 7.9% ID/g of RGD-Au-tripods had accumulated in the U87MG tumor after 24 h post-injection. A pilot mouse toxicology study confirmed that no evidence of significant acute or systemic toxicity was observed in histopathological examination. Our study suggests that Au-tripods can be reliably synthesized through stringently controlled chemical synthesis and could serve as a new generation of platform with high selectivity and sensitivity for multimodality molecular imaging.

  View details for DOI 10.1021/ja412001e

  View details for PubMedID 24495038

• Ultrasound Molecular Imaging in a Human CD276 Expression-Modulated Murine Ovarian Cancer Model. Clinical cancer research Lutz, A. M., Bachawal, S. V., Drescher, C. W., Pysz, M. A., Willmann, J. K., Gambhir, S. S. 2014; 20 (5): 1313-1322

  Abstract

  To develop a mouse ovarian cancer model that allows modulating the expression levels of human vascular targets in mouse xenograft tumors and to test whether expression of CD276 during tumor angiogenesis can be visualized by molecularly targeted ultrasound in vivo.CD276-expressing MILE SVEN 1 (MS1) mouse endothelial cells were engineered and used for coinjection with 2008 human ovarian cancer cells for subcutaneous xenograft tumor induction in 15 nude mice. Fourteen control mice were injected with 2008 cells only. After confirming their binding specificity in flow chamber cell attachment studies, anti-CD276 antibody-functionalized contrast microbubbles were used for in vivo CD276-targeted contrast-enhanced ultrasound imaging.CD276-targeted ultrasound imaging signal was significantly higher (P = 0.006) in mixed MS1/2008 tumors than in control tumors. Compared with control microbubbles, the ultrasound signal using CD276-targeted microbubbles was significantly higher (P = 0.002), and blocking with purified anti-CD276 antibody significantly decreased (P = 0.0096) the signal in mixed MS1/2008 tumors. Immunofluorescence analysis of the tumor tissue confirmed higher quantitative immunofluorescence signal in mixed MS1/2008 tumors than in control 2008 only tumors, but showed not significantly different (P = 0.54) microvessel density.Our novel small animal model allows for modulating the expression of human tumor-associated vascular endothelial imaging targets in a mouse host and these expression differences can be visualized noninvasively by ultrasound molecular imaging. The animal model can be applied to other human vascular targets and may facilitate the preclinical development of new imaging probes such as microbubbles targeted at human vascular markers not expressed in mice. Clin Cancer Res; 20(5); 1313-22. ©2014 AACR.

  View details for DOI 10.1158/1078-0432.CCR-13-1642

  View details for PubMedID 24389327

  View details for PubMedCentralID PMC3965293

• Semiconducting polymer nanoparticles as photoacoustic molecular imaging probes in living mice. Nature nanotechnology Pu, K., Shuhendler, A. J., Jokerst, J. V., Mei, J., Gambhir, S. S., Bao, Z., Rao, J. 2014; 9 (3): 233-239

  Abstract

  Photoacoustic imaging holds great promise for the visualization of physiology and pathology at the molecular level with deep tissue penetration and fine spatial resolution. To fully utilize this potential, photoacoustic molecular imaging probes have to be developed. Here, we introduce near-infrared light absorbing semiconducting polymer nanoparticles as a new class of contrast agents for photoacoustic molecular imaging. These nanoparticles can produce a stronger signal than the commonly used single-walled carbon nanotubes and gold nanorods on a per mass basis, permitting whole-body lymph-node photoacoustic mapping in living mice at a low systemic injection mass. Furthermore, the semiconducting polymer nanoparticles possess high structural flexibility, narrow photoacoustic spectral profiles and strong resistance to photodegradation and oxidation, enabling the development of the first near-infrared ratiometric photoacoustic probe for in vivo real-time imaging of reactive oxygen species-vital chemical mediators of many diseases. These results demonstrate semiconducting polymer nanoparticles to be an ideal nanoplatform for developing photoacoustic molecular probes.

  View details for DOI 10.1038/nnano.2013.302

  View details for PubMedID 24463363

  View details for PubMedCentralID PMC3947658

• Light in and sound out: emerging translational strategies for photoacoustic imaging. Cancer research Zackrisson, S., van de Ven, S. M., Gambhir, S. S. 2014; 74 (4): 979-1004

  Abstract

  Photoacoustic imaging (PAI) has the potential for real-time molecular imaging at high resolution and deep inside the tissue, using nonionizing radiation and not necessarily depending on exogenous imaging agents, making this technique very promising for a range of clinical applications. The fact that PAI systems can be made portable and compatible with existing imaging technologies favors clinical translation even more. The breadth of clinical applications in which photoacoustics could play a valuable role include: noninvasive imaging of the breast, sentinel lymph nodes, skin, thyroid, eye, prostate (transrectal), and ovaries (transvaginal); minimally invasive endoscopic imaging of gastrointestinal tract, bladder, and circulating tumor cells (in vivo flow cytometry); and intraoperative imaging for assessment of tumor margins and (lymph node) metastases. In this review, we describe the basics of PAI and its recent advances in biomedical research, followed by a discussion of strategies for clinical translation of the technique. Cancer Res; 74(4); 979-1004. ©2014 AACR.

  View details for DOI 10.1158/0008-5472.CAN-13-2387

  View details for PubMedID 24514041

  View details for PubMedCentralID PMC3944207

• Antiviral drug ganciclovir is a potent inhibitor of microglial proliferation and neuroinflammation. journal of experimental medicine Ding, Z., Mathur, V., Ho, P. P., James, M. L., Lucin, K. M., Hoehne, A., Alabsi, H., Gambhir, S. S., Steinman, L., Luo, J., Wyss-Coray, T. 2014; 211 (2): 189-198

  Abstract

  Aberrant microglial responses contribute to neuroinflammation in many neurodegenerative diseases, but no current therapies target pathogenic microglia. We discovered unexpectedly that the antiviral drug ganciclovir (GCV) inhibits the proliferation of microglia in experimental autoimmune encephalomyelitis (EAE), a mouse model for multiple sclerosis (MS), as well as in kainic acid-induced excitotoxicity. In EAE, GCV largely prevented infiltration of T lymphocytes into the central nervous system (CNS) and drastically reduced disease incidence and severity when delivered before the onset of disease. In contrast, GCV treatment had minimal effects on peripheral leukocyte distribution in EAE and did not inhibit generation of antibodies after immunization with ovalbumin. Additionally, a radiolabeled analogue of penciclovir, [(18)F]FHBG, which is similar in structure to GCV, was retained in areas of CNS inflammation in EAE, but not in naive control mice, consistent with the observed therapeutic effects. Our experiments suggest GCV may have beneficial effects in the CNS beyond its antiviral properties.

  View details for DOI 10.1084/jem.20120696

  View details for PubMedID 24493798

  View details for PubMedCentralID PMC3920559

• Evaluation of s-1 receptor radioligand 18F-FTC-146 in rats and squirrel monkeys using PET. Journal of nuclear medicine : official publication, Society of Nuclear Medicine James, M. L., Shen, B., Nielsen, C. H., Behera, D., Buckmaster, C. L., Mesangeau, C., Zavaleta, C., Vuppala, P. K., Jamalapuram, S., Avery, B. A., Lyons, D. M., McCurdy, C. R., Biswal, S., Gambhir, S. S., Chin, F. T. 2014; 55 (1): 147-153

  Abstract

  The noninvasive imaging of σ-1 receptors (S1Rs) could provide insight into their role in different diseases and lead to novel diagnostic/treatment strategies. The main objective of this study was to assess the S1R radiotracer (18)F-FTC-146 in rats. Preliminary squirrel monkey imaging and human serum/liver microsome studies were performed to gain information about the potential of (18)F-FTC-146 for eventual clinical translation.The distribution and stability of (18)F-FTC-146 in rats were assessed via PET/CT, autoradiography, γ counting, and high-performance liquid chromatography (HPLC). Preliminary PET/MRI of squirrel monkey brain was conducted along with HPLC assessment of (18)F-FTC-146 stability in monkey plasma and human serum.Biodistribution studies showed that (18)F-FTC-146 accumulated in S1R-rich rat organs, including the lungs, pancreas, spleen, and brain. Pretreatment with known S1R compounds, haloperidol, or BD1047, before radioligand administration, significantly attenuated (18)F-FTC-146 accumulation in all rat brain regions by approximately 85% (P < 0.001), suggesting radiotracer specificity for S1Rs. Similarly, PET/CT and autoradiography results demonstrated accumulation of (18)F-FTC-146 in rat brain regions known to contain S1Rs and that this uptake could be blocked by BD1047 pretreatment. Ex vivo analysis of (18)F-FTC-146 in the brain showed that only intact radiotracer was present at 15, 30, and 60 min, whereas rapid metabolism of residual (18)F-FTC-146 was observed in rat plasma. Preliminary monkey PET/MRI studies demonstrated specific accumulation of (18)F-FTC-146 in the brain (mainly in cortical structures, cerebellum, and vermis) that could be attenuated by pretreatment with haloperidol. HPLC of monkey plasma suggested radioligand metabolism, whereas (18)F-FTC-146 appeared to be stable in human serum. Finally, liver microsome studies revealed that (18)F-FTC-146 has a longer half-life in human microsomes, compared with rodents.Together, these results indicate that (18)F-FTC-146 is a promising tool for visualizing S1Rs in preclinical studies and that it has potential for mapping these sites in the human brain.

  View details for DOI 10.2967/jnumed.113.120261

  View details for PubMedID 24337599

• Imaging of Heptocellular Carcinoma Patient-Derived Xenografts Using 89Zr-Labeled Anti-Glypican-3 Monoclonal Antibody Biomaterials Yang, X., Liu, H., Sun, C. K., Natarajan, A., Hu, X., Wang, X., Allegretta, M., Guttmann, R. D., Gambhir, S. S. 2014: 6964-6971
• Pediatric Molecular Imaging Pediatric Nuclear Medicine and Molecular Imaging Daldrup-Link, H., Gambhir, S. S. edited by Treves, T. 2014; 4: 571–596
• Investigation of 6-[¹8F]-fluoromaltose as a novel PET tracer for imaging bacterial infection. PloS one Gowrishankar, G., Namavari, M., Jouannot, E. B., Hoehne, A., Reeves, R., Hardy, J., Gambhir, S. S. 2014; 9 (9)

  Abstract

  Despite advances in the field of nuclear medicine, the imaging of bacterial infections has remained a challenge. The existing reagents suffer from poor sensitivity and specificity. In this study we investigate the potential of a novel PET (positron emission tomography) tracer that overcomes these limitations.6-[¹⁸F]-fluoromaltose was synthesized. Its behavior in vitro was evaluated in bacterial and mammalian cultures. Detailed pharmacokinetic and biodistribution profiles for the tracer were obtained from a murine model.6-[¹⁸F]-fluoromaltose is taken up by multiple strains of pathogenic bacteria. It is not taken up by mammalian cancer cell lines. 6-[¹⁸F]-fluoromaltose is retained in infected muscles in a murine model of bacterial myositis. It does not accumulate in inflamed tissue.We have shown that 6-[¹⁸F]-fluoromaltose can be used to image bacterial infection in vivo with high specificity. We believe that this class of agents will have a significant impact on the clinical management of patients.

  View details for DOI 10.1371/journal.pone.0107951

  View details for PubMedID 25243851

• Tracking cellular and immune therapies in cancer. Advances in cancer research Kurtz, D. M., Gambhir, S. S. 2014; 124: 257-296

  Abstract

  The field of tumor immunology has seen an explosion of renewed interest over the last decade. With the FDA approval of new immunotherapies for prostate cancer and melanoma, as well as several exciting new drugs in clinical trials, tumor immunology is becoming an increasingly important topic in preclinical studies and patient care. However, the current methods for assessing the immune status of a patient and tumor are limited, which has led to the development of novel molecular imaging methods for assessing tumor immunology. From cell tracking for cellular therapeutics to assessing the tumor immune microenvironment, these imaging methods have the potential to further preclinical understanding of immunotherapies and potentially translate into clinically useful tests to predict and assess therapeutic response of these exciting new agents. In this review, we first discuss the recent advances in cancer immunotherapy, followed by a detailed review of the current state of molecular imaging for tumor immunology. Finally, we discuss opportunities for further development and innovation in this rapidly growing field.

  View details for DOI 10.1016/B978-0-12-411638-2.00008-2

  View details for PubMedID 25287692

• Selective uptake of single-walled carbon nanotubes by circulating monocytes for enhanced tumour delivery Nature Nanotechnology Smith, B. R., Ghosn, E. E., et al 2014: 481–87

  Abstract

  In cancer imaging, nanoparticle biodistribution is typically visualized in living subjects using 'bulk' imaging modalities such as magnetic resonance imaging, computerized tomography and whole-body fluorescence. Accordingly, nanoparticle influx is observed only macroscopically, and the mechanisms by which they target cancer remain elusive. Nanoparticles are assumed to accumulate via several targeting mechanisms, particularly extravasation (leakage into tumour). Here, we show that, in addition to conventional nanoparticle-uptake mechanisms, single-walled carbon nanotubes are almost exclusively taken up by a single immune cell subset, Ly-6C(hi) monocytes (almost 100% uptake in Ly-6C(hi) monocytes, below 3% in all other circulating cells), and delivered to the tumour in mice. We also demonstrate that a targeting ligand (RGD) conjugated to nanotubes significantly enhances the number of single-walled carbon nanotube-loaded monocytes reaching the tumour (P < 0.001, day 7 post-injection). The remarkable selectivity of this tumour-targeting mechanism demonstrates an advanced immune-based delivery strategy for enhancing specific tumour delivery with substantial penetration.

  View details for DOI 10.1038/nnano.2014.62

• Detection and quantitation of circulating tumor cell dynamics by bioluminescence imaging in an orthotopic mammary carcinoma model. PloS one Sasportas, L. S., Hori, S. S., Pratx, G., Gambhir, S. S. 2014; 9 (9): e105079

  Abstract

  Circulating tumor cells (CTCs) have been detected in the bloodstream of both early-stage and advanced cancer patients. However, very little is know about the dynamics of CTCs during cancer progression and the clinical relevance of longitudinal CTC enumeration. To address this, we developed a simple bioluminescence imaging assay to detect CTCs in mouse models of metastasis. In a 4T1 orthotopic metastatic mammary carcinoma mouse model, we demonstrated that this quantitative method offers sensitivity down to 2 CTCs in 0.1-1mL blood samples and high specificity for CTCs originating from the primary tumor, independently of their epithelial status. In this model, we simultaneously monitored blood CTC dynamics, primary tumor growth, and lung metastasis progression over the course of 24 days. Early in tumor development, we observed low numbers of CTCs in blood samples (10-15 cells/100 µL) and demonstrated that CTC dynamics correlate with viable primary tumor growth. To our knowledge, these data represent the first reported use of bioluminescence imaging to detect CTCs and quantify their dynamics in any cancer mouse model. This new assay is opening the door to the study of CTC dynamics in a variety of animal models. These studies may inform clinical decision on the appropriate timing of blood sampling and value of longitudinal CTC enumeration in cancer patients.

  View details for DOI 10.1371/journal.pone.0105079

  View details for PubMedID 25188396

  View details for PubMedCentralID PMC4154864

• Cellulose nanoparticles: Photoacoustic contrast agents that biodegrade to simple sugars Conference on Photons Plus Ultrasound: Imaging and Sensing Jokerst, J. V., Bohndiek, S. E., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2014

  View details for DOI 10.1117/12.2036256

  View details for Web of Science ID 000338768500011

• Photoacoustic Imaging of Mesenchymal Stem Cells in Living Mice via Silica-Coated Gold Nanorods Conference on Photons Plus Ultrasound: Imaging and Sensing Jokerst, J. V., Thangaraj, M., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2014

  View details for DOI 10.1117/12.2036786

  View details for Web of Science ID 000338768500035

• Tracking Cellular and Immune Therapies in Cancer EMERGING APPLICATIONS OF MOLECULAR IMAGING TO ONCOLOGY Kurtz, D. M., Gambhir, S. S. 2014; 124: 257-296

  Abstract

  The field of tumor immunology has seen an explosion of renewed interest over the last decade. With the FDA approval of new immunotherapies for prostate cancer and melanoma, as well as several exciting new drugs in clinical trials, tumor immunology is becoming an increasingly important topic in preclinical studies and patient care. However, the current methods for assessing the immune status of a patient and tumor are limited, which has led to the development of novel molecular imaging methods for assessing tumor immunology. From cell tracking for cellular therapeutics to assessing the tumor immune microenvironment, these imaging methods have the potential to further preclinical understanding of immunotherapies and potentially translate into clinically useful tests to predict and assess therapeutic response of these exciting new agents. In this review, we first discuss the recent advances in cancer immunotherapy, followed by a detailed review of the current state of molecular imaging for tumor immunology. Finally, we discuss opportunities for further development and innovation in this rapidly growing field.

  View details for DOI 10.1016/B978-0-12-411638-2.00008-2

  View details for Web of Science ID 000344511500008

• A simple model for deep tissue attenuation correction and large organ analysis of Cerenkov luminescence imaging Medical Imaging - Physics of Medical Imaging Habte, F., Natarajan, A., Paik, D. S., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2014

  View details for DOI 10.1117/12.2043879

  View details for Web of Science ID 000338775800154

• Gold nanorods combine photoacoustic and Raman imaging for detection and treatment of ovarian cancer Conference on Photons Plus Ultrasound: Imaging and Sensing Jokerst, J. V., Cole, A. J., Bohndiek, S. E., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2014

  View details for DOI 10.1117/12.2036776

  View details for Web of Science ID 000338768500134

• Detection and quantitation of circulating tumor cell dynamics by bioluminescence imaging in an orthotopic mammary carcinoma model. PloS one Sasportas, L. S., Hori, S. S., Pratx, G., Gambhir, S. S. 2014; 9 (9)

  Abstract

  Circulating tumor cells (CTCs) have been detected in the bloodstream of both early-stage and advanced cancer patients. However, very little is know about the dynamics of CTCs during cancer progression and the clinical relevance of longitudinal CTC enumeration. To address this, we developed a simple bioluminescence imaging assay to detect CTCs in mouse models of metastasis. In a 4T1 orthotopic metastatic mammary carcinoma mouse model, we demonstrated that this quantitative method offers sensitivity down to 2 CTCs in 0.1-1mL blood samples and high specificity for CTCs originating from the primary tumor, independently of their epithelial status. In this model, we simultaneously monitored blood CTC dynamics, primary tumor growth, and lung metastasis progression over the course of 24 days. Early in tumor development, we observed low numbers of CTCs in blood samples (10-15 cells/100 µL) and demonstrated that CTC dynamics correlate with viable primary tumor growth. To our knowledge, these data represent the first reported use of bioluminescence imaging to detect CTCs and quantify their dynamics in any cancer mouse model. This new assay is opening the door to the study of CTC dynamics in a variety of animal models. These studies may inform clinical decision on the appropriate timing of blood sampling and value of longitudinal CTC enumeration in cancer patients.

  View details for DOI 10.1371/journal.pone.0105079

  View details for PubMedID 25188396

  View details for PubMedCentralID PMC4154864

• Imaging circulating tumor cells in freely moving awake small animals using a miniaturized intravital microscope. PloS one Sasportas, L. S., Gambhir, S. S. 2014; 9 (1)

  Abstract

  Metastasis, the cause for 90% of cancer mortality, is a complex and poorly understood process involving the invasion of circulating tumor cells (CTCs) into blood vessels. These cells have potential prognostic value as biomarkers for early metastatic risk. But their rarity and the lack of specificity and sensitivity in measuring them render their interrogation by current techniques very challenging. How and when these cells are circulating in the blood, on their way to potentially give rise to metastasis, is a question that remains largely unanswered. In order to provide an insight into this "black box" using non-invasive imaging, we developed a novel miniature intravital microscopy (mIVM) strategy capable of real-time long-term monitoring of CTCs in awake small animals. We established an experimental 4T1-GL mouse model of metastatic breast cancer, in which tumor cells express both fluorescent and bioluminescent reporter genes to enable both single cell and whole body tumor imaging. Using mIVM, we monitored blood vessels of different diameters in awake mice in an experimental model of metastasis. Using an in-house software algorithm we developed, we demonstrated in vivo CTC enumeration and computation of CTC trajectory and speed. These data represent the first reported use we know of for a miniature mountable intravital microscopy setup for in vivo imaging of CTCs in awake animals.

  View details for DOI 10.1371/journal.pone.0086759

  View details for PubMedID 24497977

  View details for PubMedCentralID PMC3908955

• A Novel Engineered Anti-CD20 Tracer Enables Early Time PET Imaging in a Humanized Transgenic Mouse Model of B-cell Non-Hodgkins Lymphoma CLINICAL CANCER RESEARCH Natarajan, A., Hackel, B. J., Gambhir, S. S. 2013; 19 (24): 6820-6829

  Abstract

  The aim of this article was to evaluate the use of a novel engineered anti-CD20 protein based on the 10 kDa human fibronectin type 3 domain (FN3) and subsequently compare with (64)Cu-rituximab for positron emission tomography (PET) imaging of CD20.The engineered FN3(CD20) and FN3(WT) were produced in Escherichia coli cells at 2 to 5 mg/L, conjugated to DOTA, labeled with (64)Cu, and used for PET imaging of huCD20 expression in B cells. Humanized transgenic mice and subcutaneously xenografted mice each received intravenous (64)Cu-FN3(CD20) or FN3(WT) (3.7 MBq/4 μg Do-FN3 in 200 μL PBS). Control group received a blocking dose (50-fold excess) of unconjugated FN3(CD20) two hours before radiotracer injection. PET imaging was carried out at 1 to 24 hours postinjections.In vitro assay demonstrated FN3 binds CD20 with 20 nmol/L affinity on CD20-expressing cells. (64)Cu-FN3(CD20) showed clear, high-contrast visualization of huCD20-expressing B cells in the spleen of transgenic mice as early as 1 hour postinjection [38 ± 3% injected dose (ID)/g] and exhibited a spleen-to-blood ratio of 13 by 4 hours. This is higher uptake (P = 0.04) and 10-fold greater signal-to-background (P = 0.04) than the (64)Cu-rituximab antibody radiotracer. Tumor uptake (16.8 ± 1.6 vs. 5.6 ± 1.4%ID/g) and tumor:background ratios were superior for FN3CD20 relative to rituximab in xenograft studies as well.The (64)Cu-Do-FN3(CD20) radiotracer represents a novel small, high-affinity binder for imaging human CD20, which may be well suited for B-cell non-Hodgkin's lymphoma imaging in patients at early time points.

  View details for DOI 10.1158/1078-0432.CCR-13-0626

  View details for Web of Science ID 000328938700019

  View details for PubMedID 24097872

• A F-18-Labeled Saxitoxin Derivative for in Vivo PET-MR Imaging of Voltage-Gated Sodium Channel Expression Following Nerve Injury JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Hoehne, A., Behera, D., Parsons, W. H., James, M. L., Shen, B., Borgohain, P., Bodapati, D., Prabhakar, A., Gambhir, S. S., Yeomans, D. C., Biswal, S., Chin, F. T., Du Bois, J. 2013; 135 (48): 18012-18015

  Abstract

  Both chronic and neuropathic pain conditions are associated with increased expression of certain voltage-gated sodium ion channel (NaV) isoforms in peripheral sensory neurons. A method for noninvasive imaging of these channels could represent a powerful tool for investigating aberrant expression of NaV and its role in pain pathogenesis. Herein, we describe the synthesis and evaluation of a positron emission tomography (PET) radiotracer targeting NaVs, the design of which is based on the potent, NaV-selective inhibitor saxitoxin. Both autoradiography analysis of sciatic nerves excised from injured rats as well as whole animal PET-MR imaging demonstrate that a systemically administered [(18)F]-labeled saxitoxin derivative concentrates at the site of nerve injury, consistent with upregulated sodium channel expression following axotomy. This type of PET agent has potential use for serial monitoring of channel expression levels at injured nerves throughout wound healing and/or following drug treatment. Such information may be correlated with pain behavioral analyses to help shed light on the complex molecular processes that underlie pain sensation.

  View details for DOI 10.1021/ja408300e

  View details for Web of Science ID 000328100000002

  View details for PubMedID 24261833

• A (18)F-Labeled Saxitoxin Derivative for in Vivo PET-MR Imaging of Voltage-Gated Sodium Channel Expression Following Nerve Injury. Journal of the American Chemical Society Hoehne, A., Behera, D., Parsons, W. H., James, M. L., Shen, B., Borgohain, P., Bodapati, D., Prabhakar, A., Gambhir, S. S., Yeomans, D. C., Biswal, S., Chin, F. T., Bois, J. D. 2013; 135 (48): 18012-18015

  Abstract

  Both chronic and neuropathic pain conditions are associated with increased expression of certain voltage-gated sodium ion channel (NaV) isoforms in peripheral sensory neurons. A method for noninvasive imaging of these channels could represent a powerful tool for investigating aberrant expression of NaV and its role in pain pathogenesis. Herein, we describe the synthesis and evaluation of a positron emission tomography (PET) radiotracer targeting NaVs, the design of which is based on the potent, NaV-selective inhibitor saxitoxin. Both autoradiography analysis of sciatic nerves excised from injured rats as well as whole animal PET-MR imaging demonstrate that a systemically administered [(18)F]-labeled saxitoxin derivative concentrates at the site of nerve injury, consistent with upregulated sodium channel expression following axotomy. This type of PET agent has potential use for serial monitoring of channel expression levels at injured nerves throughout wound healing and/or following drug treatment. Such information may be correlated with pain behavioral analyses to help shed light on the complex molecular processes that underlie pain sensation.

  View details for DOI 10.1021/ja408300e

  View details for PubMedID 24261833

• Combined 18F-fluoride and 18F-FDG PET/CT: a response based on actual data from prospective studies. European journal of nuclear medicine and molecular imaging Iagaru, A., Mosci, C., Dick, D. W., Sathekge, M., Lapa, P., de Lima, J. M., Gambhir, S. S. 2013; 40 (12): 1922-1924

  View details for DOI 10.1007/s00259-013-2556-y

  View details for PubMedID 24057457

• Single-cell photonic nanocavity probes. Nano letters Shambat, G., Kothapalli, S., Provine, J., Sarmiento, T., Harris, J., Gambhir, S. S., Vuckovic, J. 2013; 13 (11): 4999-5005

  Abstract

  In this report, we demonstrate for the first time photonic nanocavities operating inside single biological cells. Here we develop a nanobeam photonic crystal (PC) cavity as an advanced cellular nanoprobe, active in nature, and configurable to provide a multitude of actions for both intracellular sensing and control. Our semiconductor nanocavity probes emit photoluminescence (PL) from embedded quantum dots (QD) and sustain high quality resonant photonic modes inside cells. The probes are shown to be minimally cytotoxic to cells from viability studies, and the beams can be loaded in cells and tracked for days at a time, with cells undergoing regular division with the beams. We present in vitro label-free protein sensing with our probes to detect streptavidin as a path towards real-time biomarker and biomolecule detection inside single cells. The results of this work will enable new areas of research merging the strengths of photonic nanocavities with fundamental cell biology.

  View details for DOI 10.1021/nl304602d

  View details for PubMedID 23387382

• Nanooncology: The Future of Cancer Diagnosis and Therapy CA-A CANCER JOURNAL FOR CLINICIANS Thakor, A. S., Gambhir, S. S. 2013; 63 (6): 395-418

  Abstract

  In recent years, there has been an unprecedented expansion in the field of nanomedicine with the development of new nanoparticles for the diagnosis and treatment of cancer. Nanoparticles have unique biological properties given their small size and large surface area-to-volume ratio, which allows them to bind, absorb, and carry compounds such as small molecule drugs, DNA, RNA, proteins, and probes with high efficiency. Their tunable size, shape, and surface characteristics also enable them to have high stability, high carrier capacity, the ability to incorporate both hydrophilic and hydrophobic substances and compatibility with different administration routes, thereby making them highly attractive in many aspects of oncology. This review article will discuss how nanoparticles are able to function as carriers for chemotherapeutic drugs to increase their therapeutic index; how they can function as therapeutic agents in photodynamic, gene, and thermal therapy; and how nanoparticles can be used as molecular imaging agents to detect and monitor cancer progression.

  View details for DOI 10.3322/caac.21199

  View details for PubMedID 24114523

• Integrin-Targeted Molecular Imaging of Experimental Abdominal Aortic Aneurysms by 18F-labeled Arg-Gly-Asp Positron-Emission Tomography. Circulation. Cardiovascular imaging Kitagawa, T., Kosuge, H., Chang, E., James, M. L., Yamamoto, T., Shen, B., Chin, F. T., Gambhir, S. S., Dalman, R. L., McConnell, M. V. 2013; 6 (6): 950-956

  Abstract

  Background- Both inflammation and neoangiogenesis contribute to abdominal aortic aneurysm (AAA) disease. Arg-Gly-Asp-based molecular imaging has been shown to detect the integrin αvβ3. We studied a clinical dimeric (18)F-labeled Arg-Gly-Asp positron-emission tomography (PET) agent ((18)F-FPPRGD2) for molecular imaging of experimental AAAs. Methods and Results- Murine AAAs were induced in Apo-E-deficient mice by angiotensin II infusion, with monitoring of aortic diameter on ultrasound. AAA (n=10) and saline-infused control mice (n=7) were injected intravenously with (18)F-FPPRGD2, as well as an intravascular computed tomography contrast agent, then scanned using a small-animal PET/computed tomography scanner. Aortic uptake of (18)F-FPPRGD2 was quantified by percentage-injected dose per gram and target-to-=0.003; median target-to-=0.0008). Ex vivo autoradiography demonstrated high uptake of (18)F-FPPRGD2 into the AAA wall, with immunohistochemistry showing substantial cluster of differentiation (CD)-11b(+) macrophages and CD-31(+) neovessels. Target-to-=-0.29, P=0.41) but did strongly correlate with both mural macrophage density (r=0.79, P=0.007) and neovessel counts (r=0.87, P=0.001) on immunohistochemistry. Conclusions- PET imaging of experimental AAAs using (18)F-FPPRGD2 detects biologically active disease, correlating to the degree of vascular inflammation and neoangiogenesis. This may provide a clinically translatable molecular imaging approach to characterize AAA biology to predict risk beyond size alone.

  View details for DOI 10.1161/CIRCIMAGING.113.000234

  View details for PubMedID 23995363

• Preclinical Efficacy of the Anti-Hepatocyte Growth Factor Antibody Ficlatuzumab in a Mouse Brain Orthotopic Glioma Model Evaluated by Bioluminescence, PET, and MRI CLINICAL CANCER RESEARCH Mittra, E. S., Fan-Minogue, H., Lin, F. I., Karamchandani, J., Sriram, V., Han, M., Gambhir, S. S. 2013; 19 (20): 5711-5721

  Abstract

  Ficlatuzumab is a novel therapeutic agent targeting the hepatocyte growth factor (HGF)/c-MET pathway. We summarize extensive preclinical work using this agent in a mouse brain orthotopic model of glioblastoma.Sequential experiments were done using eight- to nine-week-old nude mice injected with 3 × 10(5) U87 MG (glioblastoma) cells into the brain. Evaluation of ficlatuzumab dose response for this brain tumor model and comparison of its response to ficlatuzumab and to temozolamide were conducted first. Subsequently, various small-animal imaging modalities, including bioluminescence imaging (BLI), positron emission tomography (PET), and MRI, were used with a U87 MG-Luc 2 stable cell line, with and without the use of ficlatuzumab, to evaluate the ability to noninvasively assess tumor growth and response to therapy. ANOVA was conducted to evaluate for significant differences in the response.There was a survival benefit with ficlatuzumab alone or in combination with temozolamide. BLI was more sensitive than PET in detecting tumor cells. Fluoro-D-thymidine (FLT) PET provided a better signal-to-background ratio than 2[(18)F]fluoro-2-deoxy-d-glucose (FDG) PET. In addition, both BLI and FLT PET showed significant changes over time in the control group as well as with response to therapy. MRI does not disclose any time-dependent change. Also, the MRI results showed a temporal delay in comparison to the BLI and FLT PET findings, showing similar results one drug cycle later.Targeting the HGF/c-MET pathway with the novel agent ficlatuzumab appears promising for the treatment of glioblastoma. Various clinically applicable imaging modalities including FLT, PET, and MRI provide reliable ways of assessing tumor growth and response to therapy. Given the clinical applicability of these findings, future studies on patients with glioblastoma may be appropriate.

  View details for DOI 10.1158/1078-0432.CCR-12-1015

  View details for Web of Science ID 000325797600019

  View details for PubMedID 23983258

• A scanning transmission electron microscopy approach to analyzing large volumes of tissue to detect nanoparticles. Microscopy and microanalysis Kempen, P. J., Thakor, A. S., Zavaleta, C., Gambhir, S. S., Sinclair, R. 2013; 19 (5): 1290-1297

  Abstract

  The use of nanoparticles for the diagnosis and treatment of cancer requires the complete characterization of their toxicity, including accurately locating them within biological tissues. Owing to their size, traditional light microscopy techniques are unable to resolve them. Transmission electron microscopy provides the necessary spatial resolution to image individual nanoparticles in tissue, but is severely limited by the very small analysis volume, usually on the order of tens of cubic microns. In this work, we developed a scanning transmission electron microscopy (STEM) approach to analyze large volumes of tissue for the presence of polyethylene glycol-coated Raman-active-silica-gold-nanoparticles (PEG-R-Si-Au-NPs). This approach utilizes the simultaneous bright and dark field imaging capabilities of STEM along with careful control of the image contrast settings to readily identify PEG-R-Si-Au-NPs in mouse liver tissue without the need for additional time-consuming analytical characterization. We utilized this technique to analyze 243,000 μm3 of mouse liver tissue for the presence of PEG-R-Si-Au-NPs. Nanoparticles injected into the mice intravenously via the tail vein accumulated in the liver, whereas those injected intrarectally did not, indicating that they remain in the colon and do not pass through the colon wall into the systemic circulation.

  View details for DOI 10.1017/S143192761300192X

  View details for PubMedID 23803218

• Noninvasive imaging of hypoxia-inducible factor-1a gene therapy for myocardial ischemia. Human gene therapy methods Chen, I. Y., Gheysens, O., Li, Z., Rasooly, J. A., Wang, Q., Paulmurugan, R., Rosenberg, J., Rodriguez-Porcel, M., Willmann, J. K., Wang, D. S., Contag, C. H., Robbins, R. C., Wu, J. C., Gambhir, S. S. 2013; 24 (5): 279-288

  Abstract

  Abstract Hypoxia-inducible factor-1 alpha (HIF-1α) gene therapy holds great promise for the treatment of myocardial ischemia. Both preclinical and clinical evaluations of this therapy are underway and can benefit from a vector strategy that allows noninvasive assessment of HIF-1α expression as an objective measure of gene delivery. We have developed a novel bidirectional plasmid vector (pcTnT-HIF-1α-VP2-TSTA-fluc), which employs the cardiac troponin T (cTnT) promoter in conjunction with a two-step transcriptional amplification (TSTA) system to drive the linked expression of a recombinant HIF-1α gene (HIF-1α-VP2) and the firefly luciferase gene (fluc). The firefly luciferase (FLuc) activity serves as a surrogate for HIF-1α-VP2 expression, and can be noninvasively assessed in mice using bioluminescence imaging after vector delivery. Transfection of cultured HL-1 cardiomyocytes with pcTnT-HIF-1α-VP2-TSTA-fluc led to a strong correlation between FLuc and HIF-1α-dependent vascular endothelial growth factor expression (r(2)=0.88). Intramyocardial delivery of pcTnT-HIF-1α-VP2-TSTA-fluc into infarcted mouse myocardium led to persistent HIF-1α-VP2 expression for 4 weeks, even though it improved neither CD31+ microvessel density nor echocardiographically determined left ventricular systolic function. These results lend support to recent findings of suboptimal efficacy associated with plasmid-mediated HIF-1α therapy. The imaging techniques developed herein should be useful for further optimizing HIF-1α-VP2 therapy in preclinical models of myocardial ischemia.

  View details for DOI 10.1089/hgtb.2013.028

  View details for PubMedID 23937265

• Pilot Prospective Evaluation of Early Response to Bevacizumab Treatment Using the Novel PET/CT Radiopharmaceutical 18F FPPRGD2 Iagaru, A., Mosci, C., Davidzon, G., Kumar, M., Shen, B., Chin, F., Gambhir, S. S. SPRINGER. 2013: S185

  View details for Web of Science ID 000325853400290

• Noninvasive imaging of hypoxia-inducible factor-1a gene therapy for myocardial ischemia. Human gene therapy methods Chen, I. Y., Gheysens, O., Li, Z., Rasooly, J. A., Wang, Q., Paulmurugan, R., Rosenberg, J., Rodriguez-Porcel, M., Willmann, J. K., Wang, D. S., Contag, C. H., Robbins, R. C., Wu, J. C., Gambhir, S. S. 2013; 24 (5): 279-288

  View details for DOI 10.1089/hgtb.2013.028

  View details for PubMedID 23937265

• MicroRNA-regulated non-viral vectors with improved tumor specificity in an orthotopic rat model of hepatocellular carcinoma GENE THERAPY Ronald, J. A., Katzenberg, R., Nielsen, C. H., Jae, H. J., Hofmann, L. V., Gambhir, S. S. 2013; 20 (10): 1006-1013

  Abstract

  In hepatocellular carcinoma (HCC), tumor specificity of gene therapy is of utmost importance to preserve liver function. MicroRNAs (miRNAs) are powerful negative regulators of gene expression and many are downregulated in human HCC. We identified seven miRNAs that are also downregulated in tumors in a rat hepatoma model (P<0.05) and attempted to improve tumor specificity by constructing a panel of luciferase-expressing vectors containing binding sites for these miRNAs. Attenuation of luciferase expression by the corresponding miRNAs was confirmed across various cell lines and in mouse liver. We then tested our vectors in tumor-bearing rats and identified two miRNAs, miR-26a and miR-122, that significantly decreased expression in liver compared with the control vector (6.40 and 0.26%, respectively; P<0.05). In tumor, miR-122 had a nonsignificant trend towards decreased (∼50%) expression, whereas miR-26 had no significant effect on tumor expression. To our knowledge, this is the first work using differentially expressed miRNAs to de-target transgene expression in an orthotopic hepatoma model and to identify miR-26a, in addition to miR-122, for de-targeting liver. Considering the heterogeneity of miRNA expression in human HCC, this information will be important in guiding development of more personalized vectors for the treatment of this devastating disease.Gene Therapy advance online publication, 30 May 2013; doi:10.1038/gt.2013.24.

  View details for DOI 10.1038/gt.2013.24

  View details for Web of Science ID 000325633500006

  View details for PubMedID 23719066

• A c-Myc Activation Sensor-Based High-Throughput Drug Screening Identifies an Antineoplastic Effect of Nitazoxanide. Molecular cancer therapeutics Fan-Minogue, H., Bodapati, S., Solow-Cordero, D., Fan, A., Paulmurugan, R., Massoud, T. F., Felsher, D. W., Gambhir, S. S. 2013; 12 (9): 1896-1905

  Abstract

  Deregulation of c-Myc plays a central role in the tumorigenesis of many human cancers. Yet, the development of drugs regulating c-Myc activity has been challenging. To facilitate the identification of c-Myc inhibitors, we developed a molecular imaging sensor based high throughput-screening (HTS) system. This system uses a cell-based assay to detect c-Myc activation in a HTS format, which is established from a pure clone of a stable breast cancer cell line that constitutively expresses a c-Myc activation sensor. Optimization of the assay performance in the HTS format resulted in uniform and robust signals at the baseline. Using this system, we performed a quantitative HTS against approximately 5,000 existing bioactive compounds from five different libraries. Thirty-nine potential hits were identified, including currently known c-Myc inhibitors. There are a few among the top potent hits that are not known for anti-c-Myc activity. One of these hits is nitazoxanide (NTZ), a thiazolide for treating human protozoal infections. Validation of NTZ in different cancer cell lines revealed a high potency for c-Myc inhibition with IC50 ranging between 10 - 500nM. Oral administration of NTZ in breast cancer xenograft mouse models significantly suppressed tumor growth by inhibition of c-Myc and induction of apoptosis. These findings suggest a potential of NTZ to be repurposed as a new anti-tumor agent for inhibition of c-Myc associated neoplasia. Our work also demonstrated the unique advantage of molecular imaging in accelerating discovery of drugs for c-Myc targeted cancer therapy.

  View details for DOI 10.1158/1535-7163.MCT-12-1243

  View details for PubMedID 23825064

• Target-Specific Molecular Imaging with Cystine Knot Peptides Kimura, R. H., Gambhir, S. S. SPRINGER WIEN. 2013: 608

  View details for Web of Science ID 000323212600159

• High-sensitivity, real-time, ratiometric imaging of surface-enhanced Raman scattering nanoparticles with a clinically translatable Raman endoscope device. Journal of biomedical optics Garai, E., Sensarn, S., Zavaleta, C. L., Van de Sompel, D., Loewke, N. O., Mandella, M. J., Gambhir, S. S., Contag, C. H. 2013; 18 (9): 096008-?

  Abstract

  ABSTRACT. Topical application and quantification of targeted, surface-enhanced Raman scattering (SERS) nanoparticles offer a new technique that has the potential for early detection of epithelial cancers of hollow organs. Although less toxic than intravenous delivery, the additional washing required to remove unbound nanoparticles cannot necessarily eliminate nonspecific pooling. Therefore, we developed a real-time, ratiometric imaging technique to determine the relative concentrations of at least two spectrally unique nanoparticle types, where one serves as a nontargeted control. This approach improves the specific detection of bound, targeted nanoparticles by adjusting for working distance and for any nonspecific accumulation following washing. We engineered hardware and software to acquire SERS signals and ratios in real time and display them via a graphical user interface. We report quantitative, ratiometric imaging with nanoparticles at pM and sub-pM concentrations and at varying working distances, up to 50 mm. Additionally, we discuss optimization of a Raman endoscope by evaluating the effects of lens material and fiber coating on background noise, and theoretically modeling and simulating collection efficiency at various working distances. This work will enable the development of a clinically translatable, noncontact Raman endoscope capable of rapidly scanning large, topographically complex tissue surfaces for small and otherwise hard to detect lesions.

  View details for DOI 10.1117/1.JBO.18.9.096008

  View details for PubMedID 24008818

  View details for PubMedCentralID PMC3763230

• Evaluation of Zr-89-rituximab Tracer by Cerenkov Luminescence Imaging and Correlation with PET in a Humanized Transgenic Mouse Model to Image NHL MOLECULAR IMAGING AND BIOLOGY Natarajan, A., Habte, F., Liu, H., Sathirachinda, A., Hu, X., Cheng, Z., Nagamine, C. M., Gambhir, S. S. 2013; 15 (4): 468-475

  Abstract

  PURPOSE: This research aimed to study the use of Cerenkov luminescence imaging (CLI) for non-Hodgkin's lymphoma (NHL) using (89)Zr-rituximab positron emission tomography (PET) tracer with a humanized transgenic mouse model that expresses human CD20 and the correlation of CLI with PET. PROCEDURES: Zr-rituximab (2.6 MBq) was tail vein-injected into transgenic mice that express the human CD20 on their B cells (huCD20TM). One group (n = 3) received 2 mg/kg pre-dose (blocking) of cold rituximab 2 h prior to tracer; a second group (n = 3) had no pre-dose (non-blocking). CLI was performed using a cooled charge-coupled device optical imager. We also performed PET imaging and ex vivo studies in order to confirm the in vivo CLI results. At each time point (4, 24, 48, 72, and 96 h), two groups of mice were imaged in vivo and ex vivo with CLI and PET, and at 96 h, organs were measured by gamma counter. RESULTS: huCD20 transgenic mice injected with (89)Zr-rituximab demonstrated a high-contrast CLI image compared to mice blocked with a cold dose. At various time points of 4-96 h post-radiotracer injection, the in vivo CLI signal intensity showed specific uptake in the spleen where B cells reside and, hence, the huCD20 biomarker is present at very high levels. The time-activity curve of dose decay-corrected CLI intensity and percent injected dose per gram of tissue of PET uptake in the spleen were increased over the time period (4-96 h). At 96 h, the (89)Zr-rituximab uptake ratio (non-blocking vs blocking) counted (mean ± standard deviation) for the spleen was 1.5 ± 0.6 for CLI and 1.9 ± 0.3 for PET. Furthermore, spleen uptake measurements (non-blocking and blocking of all time points) of CLI vs PET showed good correlation (R (2) = 0.85 and slope = 0.576), which also confirmed the corresponding correlations parameter value (R (2) = 0.834 and slope = 0.47) obtained for ex vivo measurements. CONCLUSIONS: CLI and PET of huCD20 transgenic mice injected with (89)Zr-rituximab demonstrated that the tracer was able to target huCD20-expressing B cells. The in vivo and ex vivo tracer uptake corresponding to the CLI radiance intensity from the spleen is in good agreement with PET. In this report, we have validated the use of CLI with PET for NHL imaging in huCD20TM.

  View details for DOI 10.1007/s11307-013-0624-0

  View details for Web of Science ID 000321972500014

  View details for PubMedID 23471750

• Molecular imaging with surface-enhanced Raman spectroscopy nanoparticle reporters. MRS bulletin Jokerst, J. V., Pohling, C., Gambhir, S. S. 2013; 38 (8)

  Abstract

  Molecular imaging scans cellular and molecular targets in living subjects through the introduction of imaging agents that bind to these targets and report their presence through a measurable signal. The picomolar sensitivity, signal stability, and high multiplexing capacity of Raman spectroscopy satisfies important needs within the field of molecular imaging, and several groups now utilize Raman and surface-enhanced Raman spectroscopy to image molecular targets in small animal models of human disease. This article details the role of Raman spectroscopy in molecular imaging, describes some substrates and imaging agents used in animal models, and illustrates some examples.

  View details for DOI 10.1557/mrs.2013.157

  View details for PubMedID 24293809

  View details for PubMedCentralID PMC3841987

• Molecular imaging with surface-enhanced Raman spectroscopy nanoparticle reporters MRS BULLETIN Jokerst, J. V., Pohling, C., Gambhir, S. S. 2013; 38 (8): 625-630

  View details for DOI 10.1557/mrs.2013.157

  View details for Web of Science ID 000322918100014

• Real-time, continuous, fluorescence sensing in a freely-moving subject with an implanted hybrid VCSEL/CMOS biosensor BIOMEDICAL OPTICS EXPRESS O'Sullivan, T. D., Heitz, R. T., Parashurama, N., Barkin, D. B., Wooley, B. A., Gambhir, S. S., Harris, J. S., Levi, O. 2013; 4 (8): 1332-1341

  Abstract

  Performance improvements in instrumentation for optical imaging have contributed greatly to molecular imaging in living subjects. In order to advance molecular imaging in freely moving, untethered subjects, we designed a miniature vertical-cavity surface-emitting laser (VCSEL)-based biosensor measuring 1cm(3) and weighing 0.7g that accurately detects both fluorophore and tumor-targeted molecular probes in small animals. We integrated a critical enabling component, a complementary metal-oxide semiconductor (CMOS) read-out integrated circuit, which digitized the fluorescence signal to achieve autofluorescence-limited sensitivity. After surgical implantation of the lightweight sensor for two weeks, we obtained continuous and dynamic fluorophore measurements while the subject was un-anesthetized and mobile. The technology demonstrated here represents a critical step in the path toward untethered optical sensing using an integrated optoelectronic implant.

  View details for DOI 10.1364/BOE.4.001332

  View details for Web of Science ID 000322618900008

  View details for PubMedCentralID PMC3756575

• Imaging Tumor Angiogenesis: The Road to Clinical Utility AMERICAN JOURNAL OF ROENTGENOLOGY Iagaru, A., Gambhir, S. S. 2013; 201 (2): W183-W191

  Abstract

  OBJECTIVE. Tumor growth and progression require the formation of new blood vessels from preexisting vasculature, a process called angiogenesis. The ability to noninvasively visualize angiogenesis may provide new opportunities to more appropriately select patients for antiangiogenesis treatment and to monitor treatment efficacy. CONCLUSION. The superior molecular sensitivity of PET and the lack of radiation from MRI and contrast-enhanced ultrasound put these techniques at the forefront of clinical translation.

  View details for DOI 10.2214/AJR.12.8568

  View details for Web of Science ID 000322225400003

  View details for PubMedID 23883233

• Activatable oligomerizable imaging agents for photoacoustic imaging of furin-like activity in living subjects. Journal of the American Chemical Society Dragulescu-Andrasi, A., Kothapalli, S., Tikhomirov, G. A., Rao, J., Gambhir, S. S. 2013; 135 (30): 11015-11022

  Abstract

  Photoacoustic (PA) imaging is continuing to be applied for physiological imaging and more recently for molecular imaging of living subjects. Owing to its high spatial resolution in deep tissues, PA imaging holds great potential for biomedical applications and molecular diagnostics. There is however a lack of probes for targeted PA imaging, especially in the area of enzyme-activatable probes. Here we introduce a molecular probe, which upon proteolytic processing is retained at the site of enzyme activity and provides PA contrast. The probe oligomerizes via a condensation reaction and accumulates in cells and tumors that express the protease. We demonstrate that this probe reports furin and furin-like activity in cells and tumor models by generating a significantly higher PA signal relative to furin-deficient and nontarget controls. This probe could report enzyme activity in living subjects at depths significantly greater than fluorescence imaging probes and has potential for molecular imaging in deep tumors.

  View details for DOI 10.1021/ja4010078

  View details for PubMedID 23859847

• A small animal Raman instrument for rapid, wide-area, spectroscopic imaging PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Bohndiek, S. E., Wagadarikar, A., Zavaleta, C. L., Van de Sompel, D., Garai, E., Jokerst, J. V., Yazdanfar, S., Gambhir, S. S. 2013; 110 (30): 12408-12413

  Abstract

  Raman spectroscopy, amplified by surface enhanced Raman scattering (SERS) nanoparticles, is a molecular imaging modality with ultra-high sensitivity and the unique ability to multiplex readouts from different molecular targets using a single wavelength of excitation. This approach holds exciting prospects for a range of applications in medicine, including identification and characterization of malignancy during endoscopy and intraoperative image guidance of surgical resection. The development of Raman molecular imaging with SERS nanoparticles is presently limited by long acquisition times, poor spatial resolution, small field of view, and difficulty in animal handling with existing Raman spectroscopy instruments. Our goal is to overcome these limitations by designing a bespoke instrument for Raman molecular imaging in small animals. Here, we present a unique and dedicated small-animal Raman imaging instrument that enables rapid, high-spatial resolution, spectroscopic imaging over a wide field of view (> 6 cm(2)), with simplified animal handling. Imaging of SERS nanoparticles in small animals demonstrated that this small animal Raman imaging system can detect multiplexed SERS signals in both superficial and deep tissue locations at least an order of magnitude faster than existing systems without compromising sensitivity.

  View details for DOI 10.1073/pnas.1301379110

  View details for Web of Science ID 000322112300062

  View details for PubMedID 23821752

• An Observational Study of Circulating Tumor Cells and F-18-FDG PET Uptake in Patients with Treatment-Naive Non-Small Cell Lung Cancer PLOS ONE Nair, V. S., Keu, K. V., Luttgen, M. S., Kolatkar, A., Vasanawala, M., Kuschner, W., Bethel, K., Iagaru, A. H., Hoh, C., Shrager, J. B., Loo, B. W., Bazhenova, L., Nieva, J., Gambhir, S. S., Kuhn, P. 2013; 8 (7)

  Abstract

  We investigated the relationship of circulating tumor cells (CTCs) in non-small cell lung cancer (NSCLC) with tumor glucose metabolism as defined by (18)F-fluorodeoxyglucose (FDG) uptake since both have been associated with patient prognosis.We performed a retrospective screen of patients at four medical centers who underwent FDG PET-CT imaging and phlebotomy prior to a therapeutic intervention for NSCLC. We used an Epithelial Cell Adhesion Molecule (EpCAM) independent fluid biopsy based on cell morphology for CTC detection and enumeration (defined here as High Definition CTCs or "HD-CTCs"). We then correlated HD-CTCs with quantitative FDG uptake image data calibrated across centers in a cross-sectional analysis.We assessed seventy-one NSCLC patients whose median tumor size was 2.8 cm (interquartile range, IQR, 2.0-3.6) and median maximum standardized uptake value (SUVmax) was 7.2 (IQR 3.7-15.5). More than 2 HD-CTCs were detected in 63% of patients, whether across all stages (45 of 71) or in stage I disease (27 of 43). HD-CTCs were weakly correlated with partial volume corrected tumor SUVmax (r = 0.27, p-value = 0.03) and not correlated with tumor diameter (r = 0.07; p-value = 0.60). For a given partial volume corrected SUVmax or tumor diameter there was a wide range of detected HD-CTCs in circulation for both early and late stage disease.CTCs are detected frequently in early-stage NSCLC using a non-EpCAM mediated approach with a wide range noted for a given level of FDG uptake or tumor size. Integrating potentially complementary biomarkers like these with traditional patient data may eventually enhance our understanding of clinical, in vivo tumor biology in the early stages of this deadly disease.

  View details for DOI 10.1371/journal.pone.0067733

  View details for Web of Science ID 000321425300025

  View details for PubMedID 23861795

  View details for PubMedCentralID PMC3702496

• Pilot prospective evaluation of 99mTc-MDP scintigraphy, 18F NaF PET/CT, 18F FDG PET/CT and whole-body MRI for detection of skeletal metastases. Clinical nuclear medicine Iagaru, A., Young, P., Mittra, E., Dick, D. W., Herfkens, R., Gambhir, S. S. 2013; 38 (7): e290-6

  Abstract

  The aim of this study was to compare Tc-MDP bone scanning, F NaF PET/CT, F FDG PET/CT, and whole-body MRI (WBMRI) for detection of known osseous metastases.This prospective pilot trial (September 2007-April 2009) enrolled 10 participants (5 men, 5 women, 47-81 years old) diagnosed with cancer and known osseous metastases. F NaF PET/CT, F FDG PET/CT, and WBMRI were performed within 1 month for each participant.The image quality and evaluation of extent of disease were superior by F NaF PET/CT compared to Tc-MDP scintigraphy in all patients with skeletal lesions and compared to F FDG PET/CT in 3 of the patients with skeletal metastases. F NaF PET/CT showed osseous metastases where F FDG PET/CT was negative in another 3 participants. Extraskeletal metastases were identified by F FDG PET/CT in 6 participants. WBMRI with the combination of iterative decomposition of water and fat with echo asymmetry and least-squares estimation, short tau inversion recovery, and diffusion-weighted imaging pulse sequences showed fewer lesions than F NaF PET/CT in 5 patients, same number of lesions in 2 patients, and more lesions in 1 patient. WBMRI showed fewer lesions than F FDG in 3 patients and same lesions in 6 patients.Our pilot phase prospective trial demonstrated superior image quality and evaluation of skeletal disease extent with F NaF PET/CT compared to Tc-MDP scintigraphy and F FDG PET/CT, as well as the feasibility of multisequence WBMRI. In addition, F FDG PET/CT provided valuable soft-tissue information that can change disease management. Further evaluation of these findings using the recently introduced PET/MRI scanners is warranted.

  View details for DOI 10.1097/RLU.0b013e3182815f64

  View details for PubMedID 23455520

• F-18-Fluorobenzoate-Labeled Cystine Knot Peptides for PET Imaging of Integrin alpha(v)beta(6) JOURNAL OF NUCLEAR MEDICINE Hackel, B. J., Kimura, R. H., Miao, Z., Liu, H., Sathirachinda, A., Cheng, Z., Chin, F. T., Gambhir, S. S. 2013; 54 (7): 1101-1105

  Abstract

  Integrin αvβ6 is a cell surface receptor minimally expressed by healthy tissue but elevated in lung, colon, skin, ovarian, cervical, and pancreatic cancers. A molecular PET agent for integrin αvβ6 could provide significant clinical utility by facilitating both cancer staging and treatment monitoring to more rapidly identify an effective therapeutic approach. METHODS: Here, we evaluated 2 cystine knot peptides, R01 and S02, previously engineered with a 3-6 nM affinity for integrin αvβ6, for (18)F radiolabeling and PET imaging of BxPC3 pancreatic adenocarcinoma xenografts in mice. Cystine knot peptides were labeled with N-succinimidyl-4-(18)F-fluorobenzoate and evaluated for binding affinity and serum stability. Peptides conjugated with (18)F-fluorobenzoate (2-3 MBq) were injected via the tail vein into nude mice xenografted with BxPC3 (integrin αvβ6-positive) or 293 (integrin αvβ6-negative) tumors. Small-animal PET scans were acquired at 0.5, 1, and 2 h after injection. Ex vivo γ-counting of dissected tissues was performed at 0.5 and 2 h. RESULTS: (18)F-fluorobenzoate peptides were produced in 93% ((18)F-fluorobenzoate-R01) and 99% ((18)F-fluorobenzoate-S02) purity. (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02 had affinities of 1.1 ± 0.2 and 0.7 ± 0.4 nM, respectively, and were 87% and 94%, respectively, stable in human serum at 37°C for 2 h. (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02 exhibited 2.3 ± 0.6 and 1.3 ± 0.4 percentage injected dose per gram (%ID/g), respectively, in BxPC3 xenografted tumors at 0.5 h (n = 4-5). Target specificity was confirmed by low tumor uptake in integrin αvβ6-negative 293 tumors (1.4 ± 0.6 and 0.5 ± 0.2 %ID/g, respectively, for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02; both P < 0.05; n = 3-4) and low muscle uptake (3.1 ± 1.0 and 2.7 ± 0.4 tumor to muscle for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02, respectively). Small-animal PET data were corroborated by ex vivo γ-counting of dissected tissues, which demonstrated low uptake in nontarget tissues with only modest kidney uptake (9.2 ± 3.3 and 1.9 ± 1.2 %ID/g, respectively, at 2 h for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02; n = 8). Uptake in healthy pancreas was low (0.3% ± 0.1% for (18)F-fluorobenzoate-R01 and 0.03% ± 0.01% for (18)F-fluorobenzoate-S02; n = 8). CONCLUSION: These cystine knot peptide tracers, in particular (18)F-fluorobenzoate-R01, show translational promise for molecular imaging of integrin αvβ6 overexpression in pancreatic and other cancers.

  View details for DOI 10.2967/jnumed.112.110759

  View details for Web of Science ID 000321113500019

• 18F-fluorobenzoate-labeled cystine knot peptides for PET imaging of integrin avß6. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Hackel, B. J., Kimura, R. H., Miao, Z., Liu, H., Sathirachinda, A., Cheng, Z., Chin, F. T., Gambhir, S. S. 2013; 54 (7): 1101-1105

  Abstract

  Integrin αvβ6 is a cell surface receptor minimally expressed by healthy tissue but elevated in lung, colon, skin, ovarian, cervical, and pancreatic cancers. A molecular PET agent for integrin αvβ6 could provide significant clinical utility by facilitating both cancer staging and treatment monitoring to more rapidly identify an effective therapeutic approach. METHODS: Here, we evaluated 2 cystine knot peptides, R01 and S02, previously engineered with a 3-6 nM affinity for integrin αvβ6, for (18)F radiolabeling and PET imaging of BxPC3 pancreatic adenocarcinoma xenografts in mice. Cystine knot peptides were labeled with N-succinimidyl-4-(18)F-fluorobenzoate and evaluated for binding affinity and serum stability. Peptides conjugated with (18)F-fluorobenzoate (2-3 MBq) were injected via the tail vein into nude mice xenografted with BxPC3 (integrin αvβ6-positive) or 293 (integrin αvβ6-negative) tumors. Small-animal PET scans were acquired at 0.5, 1, and 2 h after injection. Ex vivo γ-counting of dissected tissues was performed at 0.5 and 2 h. RESULTS: (18)F-fluorobenzoate peptides were produced in 93% ((18)F-fluorobenzoate-R01) and 99% ((18)F-fluorobenzoate-S02) purity. (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02 had affinities of 1.1 ± 0.2 and 0.7 ± 0.4 nM, respectively, and were 87% and 94%, respectively, stable in human serum at 37°C for 2 h. (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02 exhibited 2.3 ± 0.6 and 1.3 ± 0.4 percentage injected dose per gram (%ID/g), respectively, in BxPC3 xenografted tumors at 0.5 h (n = 4-5). Target specificity was confirmed by low tumor uptake in integrin αvβ6-negative 293 tumors (1.4 ± 0.6 and 0.5 ± 0.2 %ID/g, respectively, for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02; both P < 0.05; n = 3-4) and low muscle uptake (3.1 ± 1.0 and 2.7 ± 0.4 tumor to muscle for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02, respectively). Small-animal PET data were corroborated by ex vivo γ-counting of dissected tissues, which demonstrated low uptake in nontarget tissues with only modest kidney uptake (9.2 ± 3.3 and 1.9 ± 1.2 %ID/g, respectively, at 2 h for (18)F-fluorobenzoate-R01 and (18)F-fluorobenzoate-S02; n = 8). Uptake in healthy pancreas was low (0.3% ± 0.1% for (18)F-fluorobenzoate-R01 and 0.03% ± 0.01% for (18)F-fluorobenzoate-S02; n = 8). CONCLUSION: These cystine knot peptide tracers, in particular (18)F-fluorobenzoate-R01, show translational promise for molecular imaging of integrin αvβ6 overexpression in pancreatic and other cancers.

  View details for DOI 10.2967/jnumed.112.110759

  View details for PubMedID 23670900

• Pilot prospective evaluation of 99mTc-MDP scintigraphy, 18F NaF PET/CT, 18F FDG PET/CT and whole-body MRI for detection of skeletal metastases. Clinical nuclear medicine Iagaru, A., Young, P., Mittra, E., Dick, D. W., Herfkens, R., Gambhir, S. S. 2013; 38 (7): e290-6

  View details for DOI 10.1097/RLU.0b013e3182815f64

  View details for PubMedID 23455520

• Noninvasive Monitoring of Oxidative Stress in Transplanted Mesenchymal Stromal Cells JACC-CARDIOVASCULAR IMAGING Psaltis, P. J., Peterson, K. M., Xu, R., Franchi, F., Witt, T., Chen, I. Y., Lerman, A., Simari, R. D., Gambhir, S. S., Rodriguez-Porcel, M. 2013; 6 (7): 795-802

  Abstract

  OBJECTIVES: The goal of this study was to validate a pathway-specific reporter gene that could be used to noninvasively image the oxidative status of progenitor cells. BACKGROUND: In cell therapy studies, reporter gene imaging plays a valuable role in the assessment of cell fate in living subjects. After myocardial injury, noxious stimuli in the host tissue confer oxidative stress to transplanted cells that may influence their survival and reparative function. METHODS: Rat mesenchymal stromal cells (MSCs) were studied for phenotypic evidence of increased oxidative stress under in vitro stress. On the basis of their up-regulation of the pro-oxidant enzyme p67(phox) subunit of nicotinamide adenine dinucleotide phosphate (NAD[P]H oxidase p67(phox)), an oxidative stress sensor was constructed, comprising the firefly luciferase (Fluc) reporter gene driven by the NAD(P)H p67(phox) promoter. MSCs cotransfected with NAD(P)H p67(phox)-Fluc and a cell viability reporter gene (cytomegalovirus-Renilla luciferase) were studied under in vitro and in vivo pro-oxidant conditions. RESULTS: After in vitro validation of the sensor during low-serum culture, transfected MSCs were transplanted into a rat model of myocardial ischemia/reperfusion (IR) and monitored by using bioluminescence imaging. Compared with sham controls (no IR), cardiac Fluc intensity was significantly higher in IR rats (3.5-fold at 6 h, 2.6-fold at 24 h, 5.4-fold at 48 h; p < 0.01), indicating increased cellular oxidative stress. This finding was corroborated by ex vivo luminometry after correcting for Renilla luciferase activity as a measure of viable MSC number (Fluc:Renilla luciferase ratio 0.011 ± 0.003 for sham vs. 0.026 ± 0.004 for IR at 48 h; p < 0.05). Furthermore, in IR animals that received MSCs preconditioned with an antioxidant agent (tempol), Fluc signal was strongly attenuated, substantiating the specificity of the oxidative stress sensor. CONCLUSIONS: Pathway-specific reporter gene imaging allows assessment of changes in the oxidative status of MSCs after delivery to ischemic myocardium, providing a template to monitor key biological interactions between transplanted cells and their host environment in living subjects.

  View details for DOI 10.1016/j.jcmg.2012.11.018

  View details for Web of Science ID 000321677300006

  View details for PubMedID 23643284

• A Raman-based endoscopic strategy for multiplexed molecular imaging. Proceedings of the National Academy of Sciences of the United States of America Zavaleta, C. L., Garai, E., Liu, J. T., Sensarn, S., Mandella, M. J., Van de Sompel, D., Friedland, S., Van Dam, J., Contag, C. H., Gambhir, S. S. 2013; 110 (25): E2288-97

  Abstract

  Endoscopic imaging is an invaluable diagnostic tool allowing minimally invasive access to tissues deep within the body. It has played a key role in screening colon cancer and is credited with preventing deaths through the detection and removal of precancerous polyps. However, conventional white-light endoscopy offers physicians structural information without the biochemical information that would be advantageous for early detection and is essential for molecular typing. To address this unmet need, we have developed a unique accessory, noncontact, fiber optic-based Raman spectroscopy device that has the potential to provide real-time, multiplexed functional information during routine endoscopy. This device is ideally suited for detection of functionalized surface-enhanced Raman scattering (SERS) nanoparticles as molecular imaging contrast agents. This device was designed for insertion through a clinical endoscope and has the potential to detect and quantify the presence of a multiplexed panel of tumor-targeting SERS nanoparticles. Characterization of the Raman instrument was performed with SERS particles on excised human tissue samples, and it has shown unsurpassed sensitivity and multiplexing capabilities, detecting 326-fM concentrations of SERS nanoparticles and unmixing 10 variations of colocalized SERS nanoparticles. Another unique feature of our noncontact Raman endoscope is that it has been designed for efficient use over a wide range of working distances from 1 to 10 mm. This is necessary to accommodate for imperfect centering during endoscopy and the nonuniform surface topology of human tissue. Using this endoscope as a key part of a multiplexed detection approach could allow endoscopists to distinguish between normal and precancerous tissues rapidly and to identify flat lesions that are otherwise missed.

  View details for DOI 10.1073/pnas.1211309110

  View details for PubMedID 23703909

  View details for PubMedCentralID PMC3690865

• Molecular Imaging of Inflammation in Inflammatory Bowel Disease with a Clinically Translatable Dual-Selectin-targeted US Contrast Agent: Comparison with FDG PET/CT in a Mouse Model. Radiology Wang, H., Machtaler, S., Bettinger, T., Lutz, A. M., Luong, R., Bussat, P., Gambhir, S. S., Tranquart, F., Tian, L., Willmann, J. K. 2013; 267 (3): 818-829

  Abstract

  Purpose: To develop and test a molecular imaging approach that uses ultrasonography (US) and a clinically translatable dual-targeted (P- and E-selectin) contrast agent (MBSelectin) in the quantification of inflammation at the molecular level and to quantitatively correlate selectin-targeted US with fluorodeoxyglucose (FDG) combined positron emission tomography (PET) and computed tomography (CT) in terms of visualization and quantification of different levels of inflammation in a murine acute colitis model. Materials and Methods: Animal studies were approved by the Institutional Administrative Panel on Laboratory Animal Care at Stanford University. MBSelectin was developed by covalently binding an analog of the naturally occurring binding ligand P-selectin glycoprotein ligand 1 fused to a human fragment crystallizable(or Fc) domain onto the lipid shell of perfluorobutane and nitrogen-containing MBs. Binding specificity of MBSelectin was assessed in vitro with a flow chamber assay and in vivo with a chemically induced acute colitis murine model. US signal was quantitatively correlated with FDG uptake at PET/CT and histologic grade. Statistical analysis was performed with the Student t test, analysis of variance, and Pearson correlation analysis. Results: MBSelectin showed strong attachment to both human and mouse P- and E-selectin compared with MBControl in vitro (P ≤ .002). In vivo, US signal was significantly increased (P < .001) in mice with acute colitis (173.8 arbitrary units [au] ± 134.8 [standard deviation]) compared with control mice (5.0 au ± 4.5). US imaging signal strongly correlated with FDG uptake on PET/CT images (ρ = 0.89, P < .001). Ex vivo analysis enabled confirmation of inflammation in mice with acute colitis and high expression levels of P- and E-selectin in mucosal capillaries (P = .014). Conclusion: US with MBSelectin specifically enables detection and quantification of inflammation in a murine acute colitis model, leveraging the natural pathway of leukocyte recruitment in inflammatory tissue. US imaging with MBSelectin correlates well with FDG uptake at PET/CT imaging. © RSNA, 2013 Supplemental material: http://radiology.rsna.org/lookup/suppl/doi:10.1148/radiol.13122509/-/DC1.

  View details for DOI 10.1148/radiol.13122509

  View details for PubMedID 23371306

• A comparison of noise models in a hybrid reference spectrum and principal components analysis algorithm for Raman spectroscopy JOURNAL OF RAMAN SPECTROSCOPY Van de Sompel, D., Garai, E., Zavaleta, C., Gambhir, S. S. 2013; 44 (6): 841-856

  View details for DOI 10.1002/jrs.4258

  View details for Web of Science ID 000319935700009

• Development and validation of an immuno-PET tracer for patient stratification and therapy monitoring of antibody-drug conjugate therapy Ilovich, O., Natarajan, A., Sathirachinda, A., Kimura, R., Srinivasan, A., Gebauer, M., Kruip, J., Carrez, C., Sassoon, I., Blanc, V., Sarkar, S. K., Gambhir, S. AMER SOC CLINICAL ONCOLOGY. 2013

  View details for Web of Science ID 000335419600416

• Advanced Characterization Techniques for Nanoparticles for Cancer Research: Applications of SEM and NanoSIMS for Locating Au Nanoparticles in Cells. Materials Research Society symposia proceedings. Materials Research Society Kempen, P. J., Hitzman, C., Sasportas, L. S., Gambhir, S. S., Sinclair, R. 2013; 1569: 157-163

  Abstract

  The ability of nano secondary ion mass spectrometry (NanoSIMS) to locate and analyze Raman active gold core nanoparticles (R-AuNPs) in a biological system is compared with the standard analysis using the scanning electron microscope (SEM). The same cell with R-AuNPs on and inside the macrophage was analyzed with both techniques to directly compare them. SEM analysis showed a large number of nanoparticles within the cell. Subsequent NanoSIMS analysis showed fewer R-AuNPs with lower spatial resolution. SEM was determined to be superior to NanoSIMS for the analysis of inorganic nanoparticles in complex biological systems.

  View details for PubMedID 25364091

• [F-18]CAIP a smart PET tracer for imaging caspase-3 induced Apoptosis Shen Bin, B., Jeon, J., Palner, M., Tong Ling, L., Felsher, D., Gambhir, S. S., Chin, F. T., Rao Jianghong, J. H. WILEY-BLACKWELL. 2013: S6–S6

  View details for Web of Science ID 000318694100007

• Evaluation of the antitumor effects of rilotumumab by PET imaging in a U-87 MG mouse xenograft model NUCLEAR MEDICINE AND BIOLOGY Rex, K., Lewis, X. Z., Sundaresan, G., Glaus, C., Silva, M. D., Radinsky, R., Burgess, T. L., Gambhir, S. S., Coxon, A. 2013; 40 (4): 458-463

  Abstract

  Dysregulation of the hepatocyte growth factor (HGF)/MET pathway has been implicated in various cancers. Rilotumumab is an investigational, fully human monoclonal antibody that binds and neutralizes HGF. The purpose of this study was to evaluate the efficacy of rilotumumab in a U-87 MG mouse xenograft tumor model using (18)F-FDG and (18)F-FLT PET.U-87 MG tumor-bearing nude mice received rilotumumab or control IgG2. In the dose response study, increasing doses of rilotumumab (10, 30, 100, 300, or 500 μg) were administered, and mice were evaluated with (18)F-FDG PET at baseline and 7 days post-treatment. In the time course study, 300 μg of rilotumumab twice per week was used for the treatment, and mice were evaluated over 7 days using (18)F-FDG and (18)F-FLT PET.In the dose response study, rilotumumab at doses of 300 and 500 μg was similarly effective against tumor growth. Treatment with 300 and 500 μg rilotumumab inhibited (18)F-FDG accumulation with significant decreases of -37% and -40% in the percent injected dose per gram of tissue (%ID/g), respectively. In the time course study, treatment with 300 μg rilotumumab inhibited (18)F-FDG and (18)F-FLT accumulation with a maximum %ID/g of -41% and -64%, respectively. No apparent differences between the use of either tracer to evaluate rilotumumab efficacy were observed.Rilotumumab inhibited (18)F-FDG and (18)F-FLT accumulation as early as 2 and 4 days after treatment, respectively, in a mouse tumor model. Further studies to evaluate (18)F-FDG PET imaging as an early tumor response marker for rilotumumab are warranted. Rilotumumab is currently being tested in patients with MET-positive, advanced gastric and gastroesophageal cancer.

  View details for DOI 10.1016/j.nucmedbio.2013.01.004

  View details for Web of Science ID 000325842800005

• Biodistribution and kinetics of 18F FPPRGD2 in cancer patients Davidzon, G., Mosci, C., Mittra, E., Shen, B., Chin, F., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2013

  View details for Web of Science ID 000209478700012

• Evaluation of the antitumor effects of rilotumumab by PET imaging in a U-87 MG mouse xenograft model. Nuclear medicine and biology Rex, K., Lewis, X. Z., Gobalakrishnan, S., Glaus, C., Silva, M. D., Radinsky, R., Burgess, T. L., Gambhir, S. S., Coxon, A. 2013; 40 (4): 458-463

  Abstract

  Dysregulation of the hepatocyte growth factor (HGF)/MET pathway has been implicated in various cancers. Rilotumumab is an investigational, fully human monoclonal antibody that binds and neutralizes HGF. The purpose of this study was to evaluate the efficacy of rilotumumab in a U-87 MG mouse xenograft tumor model using (18)F-FDG and (18)F-FLT PET.U-87 MG tumor-bearing nude mice received rilotumumab or control IgG2. In the dose response study, increasing doses of rilotumumab (10, 30, 100, 300, or 500 μg) were administered, and mice were evaluated with (18)F-FDG PET at baseline and 7 days post-treatment. In the time course study, 300 μg of rilotumumab twice per week was used for the treatment, and mice were evaluated over 7 days using (18)F-FDG and (18)F-FLT PET.In the dose response study, rilotumumab at doses of 300 and 500 μg was similarly effective against tumor growth. Treatment with 300 and 500 μg rilotumumab inhibited (18)F-FDG accumulation with significant decreases of -37% and -40% in the percent injected dose per gram of tissue (%ID/g), respectively. In the time course study, treatment with 300 μg rilotumumab inhibited (18)F-FDG and (18)F-FLT accumulation with a maximum %ID/g of -41% and -64%, respectively. No apparent differences between the use of either tracer to evaluate rilotumumab efficacy were observed.Rilotumumab inhibited (18)F-FDG and (18)F-FLT accumulation as early as 2 and 4 days after treatment, respectively, in a mouse tumor model. Further studies to evaluate (18)F-FDG PET imaging as an early tumor response marker for rilotumumab are warranted. Rilotumumab is currently being tested in patients with MET-positive, advanced gastric and gastroesophageal cancer.

  View details for DOI 10.1016/j.nucmedbio.2013.01.004

  View details for PubMedID 23454250

• Developing a non-invasive, diagnostic test for stage I non-small cell lung cancer using circulating tumor cells. Luttgen, M. S., Keu, K., Nair, V. S., Horng, G., Vasanawala, M., Kolatkar, A., Carlsson, A., Sabouri, M., Loo, B. W., Shrager, J. B., Iagaru, A., Kuschner, W., Kuhn, P., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2013

  View details for DOI 10.1158/1538-7445.AM2013-3485

  View details for Web of Science ID 000331220602190

• High-resolution, serial intravital microscopic imaging of nanoparticle delivery and targeting in a small animal tumor model NANO TODAY Smith, B. R., Zavaleta, C., Rosenberg, J., Tong, R., Ramunas, J., Liu, Z., Dai, H., Gambhir, S. S. 2013; 8 (2): 126-137

  View details for DOI 10.1016/j.nantod.2013.02.004

  View details for Web of Science ID 000319236400006

• High-resolution, serial intravital microscopic imaging of nanoparticle delivery and targeting in a small animal tumor model. Nano today Smith, B. R., Zavaleta, C., Rosenberg, J., Tong, R., Ramunas, J., Liu, Z., Dai, H., Gambhir, S. S. 2013; 8 (2)

  Abstract

  Nanoparticles are under active investigation for the detection and treatment of cancer. Yet our understanding of nanoparticle delivery to tumors is limited by our ability to observe the uptake process on its own scale in living subjects. We chose to study single-walled carbon nanotubes (SWNTs) because they exhibit among the highest levels of tumor uptake across the wide variety of available nanoparticles. We target them using RGD (arginine-glycine-aspartic acid) peptide which directs them to integrins overexpressed on tumor vasculature and on the surface of some tumor cells (e.g., U87MG as used here). We employ intravital microscopy (IVM) to quantitatively examine the spatiotemporal framework of targeted SWNT uptake in a murine tumor model. IVM provided a dynamic microscale window into nanoparticle circulation, binding to tumor blood vessels, extravasation, binding to tumor cells, and tumor retention. RGD-SWNTs bound to tumor vasculature significantly more than controls (P<0.0001). RGD-SWNTs extravasated similarly compared to control RAD-SWNTs, but post-extravasation we observed as RGD-SWNTs eventually bound to individual tumor cells significantly more than RAD-SWNTs (p<0.0001) over time. RGD-SWNTs and RAD-SWNTs displayed similar signal in tumor for a week, but over time their curves significantly diverged (p<0.001) showing increasing RGD-SWNTs relative to untargeted SWNTs. We uncovered the complex spatiotemporal interplay between these competing uptake mechanisms. Specific uptake was delimited to early (1-6 hours) and late (1-4 weeks) time-points, while non-specific uptake dominated from 6 hours to 1 week. Our analysis revealed critical, quantitative insights into the dynamic, multifaceted mechanisms implicated in ligand-targeted SWNT accumulation in tumor using real-time observation.

  View details for DOI 10.1016/j.nantod.2013.02.004

  View details for PubMedID 24273594

  View details for PubMedCentralID PMC3836612

• Intracellular Aggregation of Multimodal Silica Nanoparticles for Ultrasound-Guided Stem Cell Implantation SCIENCE TRANSLATIONAL MEDICINE Jokerst, J. V., Khademi, C., Gambhir, S. S. 2013; 5 (177)

  Abstract

  The promises of cardiac stem cell therapy have yet to be fully realized, in part because of poor survival and engraftment efficacy of implanted cells. Cells die after implantation owing to ischemia, inflammation, immune response, as well as mis-injection or implantation into fibrotic tissue. Imaging tools can help implant cells in areas of the heart most receptive to stem cell therapy and monitor the efficacy of treatment by reporting the viability, location, and number of implanted stem cells. We describe a multimodal, silica-based nanoparticle that can be used for cell sorting (fluorescence), real-time guided cell implantation ultrasound, and high-resolution, long-term monitoring by magnetic resonance imaging (MRI). The nanoparticle agent increased the ultrasound and MRI contrast of labeled human mesenchymal stem cells (hMSCs) 700 and 200% versus unlabeled cells, respectively, and allowed cell imaging in animal models for 13 days after implantation. The agent had no significant impact on hMSC cell metabolic activity, proliferation, or pluripotency, and it increased the production of many paracrine factors implicated in cardiac repair. Electron microscopy and ultrasound imaging suggest that the mechanism of action is in vivo aggregation of the 300-nm silica nanoparticles into larger silica frameworks that amplify the ultrasound backscatter. The detection limit in cardiac tissue was 250,000 hMSCs via MRI and 70,000 via ultrasound. This ultrasound-guided cell delivery and multimodal optical/ultrasound/MRI intracardiac cell-tracking platform could improve cell therapy in the clinic by minimizing misdelivery or implantation into fibrotic tissue.

  View details for DOI 10.1126/scitranslmed.3005228

  View details for Web of Science ID 000316454100004

  View details for PubMedID 23515077

• Molecular Photoacoustic Imaging of Follicular Thyroid Carcinoma CLINICAL CANCER RESEARCH Levi, J., Kothapalli, S., Bohndiek, S., Yoon, J., Dragulescu-Andrasi, A., Nielsen, C., Tisma, A., Bodapati, S., Gowrishankar, G., Yan, X., Chan, C., Starcevic, D., Gambhir, S. S. 2013; 19 (6): 1494-1502

  Abstract

  To evaluate the potential of targeted photoacoustic imaging as a noninvasive method for detection of follicular thyroid carcinoma.We determined the presence and activity of two members of matrix metalloproteinase family (MMP), MMP-2 and MMP-9, suggested as biomarkers for malignant thyroid lesions, in FTC133 thyroid tumors subcutaneously implanted in nude mice. The imaging agent used to visualize tumors was MMP-activatable photoacoustic probe, Alexa750-CXeeeeXPLGLAGrrrrrXK-BHQ3. Cleavage of the MMP-activatable agent was imaged after intratumoral and intravenous injections in living mice optically, observing the increase in Alexa750 fluorescence, and photoacoustically, using a dual-wavelength imaging method.Active forms of both MMP-2 and MMP-9 enzymes were found in FTC133 tumor homogenates, with MMP-9 detected in greater amounts. The molecular imaging agent was determined to be activated by both enzymes in vitro, with MMP-9 being more efficient in this regard. Both optical and photoacoustic imaging showed significantly higher signal in tumors of mice injected with the active agent than in tumors injected with the control, nonactivatable, agent.With the combination of high spatial resolution and signal specificity, targeted photoacoustic imaging holds great promise as a noninvasive method for early diagnosis of follicular thyroid carcinomas.

  View details for DOI 10.1158/1078-0432.CCR-12-3061

  View details for Web of Science ID 000316188900021

  View details for PubMedID 23349314

  View details for PubMedCentralID PMC3602312

• Earlier Detection of Breast Cancer with Ultrasound Molecular Imaging in a Transgenic Mouse Model CANCER RESEARCH Bachawal, S. V., Jensen, K. C., Lutz, A. M., Gambhir, S. S., Tranquart, F., Tian, L., Willmann, J. K. 2013; 73 (6): 1689-1698

  Abstract

  While there is an increasing role of ultrasound for breast cancer screening in patients with dense breast, conventional anatomical ultrasound lacks sensitivity and specificity for early breast cancer detection. In this study, we assessed the potential of ultrasound molecular imaging using clinically translatable vascular endothelial growth factor receptor type 2 (VEGFR2)-targeted microbubbles (MB(VEGFR2)) to improve the diagnostic accuracy of ultrasound in earlier detection of breast cancer and ductal carcinoma in situ (DCIS) in a transgenic mouse model [FVB/N-Tg(MMTV-PyMT)634Mul]. In vivo binding specificity studies (n = 26 tumors) showed that ultrasound imaging signal was significantly higher (P < 0.001) using MB(VEGFR2) than nontargeted microbubbles and imaging signal significantly decreased (P < 0.001) by blocking antibodies. Ultrasound molecular imaging signal significantly increased (P < 0.001) when breast tissue (n = 315 glands) progressed from normal [1.65 ± 0.17 arbitrary units (a.u.)] to hyperplasia (4.21 ± 1.16), DCIS (15.95 ± 1.31), and invasive cancer (78.1 ± 6.31) and highly correlated with ex vivo VEGFR2 expression [R(2) = 0.84; 95% confidence interval (CI), 0.72-0.91; P < 0.001]. At an imaging signal threshold of 4.6 a.u., ultrasound molecular imaging differentiated benign from malignant entities with a sensitivity of 84% (95% CI, 78-88) and specificity of 89% (95% CI, 81-94). In a prospective screening trail (n = 63 glands), diagnostic performance of detecting DCIS and breast cancer was assessed and two independent readers correctly diagnosed malignant disease in more than 95% of cases and highly agreed between each other [intraclass correlation coefficient (ICC) = 0.98; 95% CI, 97-99]. These results suggest that VEGFR2-targeted ultrasound molecular imaging allows highly accurate detection of DCIS and breast cancer in transgenic mice and may be a promising approach for early breast cancer detection in women.

  View details for DOI 10.1158/0008-5472.CAN-12-3391

  View details for Web of Science ID 000316187500006

  View details for PubMedID 23328585

  View details for PubMedCentralID PMC3602408

• An Integrated Computational/Experimental Model of Lymphoma Growth PLOS COMPUTATIONAL BIOLOGY Frieboes, H. B., Smith, B. R., Chuang, Y., Ito, K., Roettgers, A. M., Gambhir, S. S., Cristini, V. 2013; 9 (3)

  Abstract

  Non-Hodgkin's lymphoma is a disseminated, highly malignant cancer, with resistance to drug treatment based on molecular- and tissue-scale characteristics that are intricately linked. A critical element of molecular resistance has been traced to the loss of functionality in proteins such as the tumor suppressor p53. We investigate the tissue-scale physiologic effects of this loss by integrating in vivo and immunohistological data with computational modeling to study the spatiotemporal physical dynamics of lymphoma growth. We compare between drug-sensitive Eμ-myc Arf-/- and drug-resistant Eμ-myc p53-/- lymphoma cell tumors grown in live mice. Initial values for the model parameters are obtained in part by extracting values from the cellular-scale from whole-tumor histological staining of the tumor-infiltrated inguinal lymph node in vivo. We compare model-predicted tumor growth with that observed from intravital microscopy and macroscopic imaging in vivo, finding that the model is able to accurately predict lymphoma growth. A critical physical mechanism underlying drug-resistant phenotypes may be that the Eμ-myc p53-/- cells seem to pack more closely within the tumor than the Eμ-myc Arf-/- cells, thus possibly exacerbating diffusion gradients of oxygen, leading to cell quiescence and hence resistance to cell-cycle specific drugs. Tighter cell packing could also maintain steeper gradients of drug and lead to insufficient toxicity. The transport phenomena within the lymphoma may thus contribute in nontrivial, complex ways to the difference in drug sensitivity between Eμ-myc Arf-/- and Eμ-myc p53-/- tumors, beyond what might be solely expected from loss of functionality at the molecular scale. We conclude that computational modeling tightly integrated with experimental data gives insight into the dynamics of Non-Hodgkin's lymphoma and provides a platform to generate confirmable predictions of tumor growth.

  View details for DOI 10.1371/journal.pcbi.1003008

  View details for Web of Science ID 000316864200070

  View details for PubMedID 23555235

  View details for PubMedCentralID PMC3610621

• BIOENGINEERING AND REGENERATIVE MEDICINE Keeping track NATURE MATERIALS Ziv, K., Gambhir, S. S. 2013; 12 (3): 180-181

  View details for Web of Science ID 000315707200009

  View details for PubMedID 23422714

• Non-Invasive Imaging of Phosphoinositide-3-Kinase-Catalytic-Subunit-Alpha (PIK3CA) Promoter Modulation in Small Animal Models PLOS ONE Gaikwad, S. M., Gunjal, L., Junutula, A. R., Astanehe, A., Gambhir, S. S., Ray, P. 2013; 8 (2)

  Abstract

  Activation of the PI3K/Akt pathway, a critical step for survival in cancer cells is often associated with decreased sensitivity to several chemotherapeutic drugs. PIK3CA gene amplification is observed in 16-24% of epithelial ovarian cancer (EOC) patients in conjunction with p53 mutations. A 900 bp long PIK3CA promoter is shown to be negatively regulated by p53 in ovarian surface epithelial cells but the consequence of chemotherapeutic drug treatments on this promoter in ovarian cancer cells is largely unknown. We aim to study the modulation of this promoter by cisplatin using an improved fusion reporter in ovarian cancer cells and tumor xenografts by non-invasive imaging approach. A PIK3CA sensor was developed using a bi-fusion reporter from a newly constructed library of bi- and tri-fusion vectors comprising of two mutant far red fluorescent proteins (mcherry/mch and tdTomato/tdt), a mutant firefly luciferase (fluc2), and a PET reporter protein (ttk). In vivo imaging of mice implanted with 293T cells transiently expressing these bi- and tri-fusion reporters along with respective controls revealed comparable activity of each reporter in the fusion background and fluc2-tdt as the most sensitive one. Repression of the PIK3CA sensor by drugs was inversely proportional to cellular p53 level in a germline (PA1) and in an EOC (A2780) cell line but not in a p53 deficient EOC (SKOV3) cell line. Bioluminescence imaging of tumor xenografts stably expressing the PIK3CA sensor in PA1 and A2780 cells exhibited attenuating activity without any change in SKOV3 tumors expressing the PIK3CA sensor after cisplatin treatment. Sequential mutation at p53 binding sites showed gradual increase in promoter activity and decreased effects of the drugs. These newly developed PIK3CA-fluc2-tdt and the mutant reporter sensors thus would be extremely useful for screening new drugs and for functional assessment of PIK3CA expression from intact cells to living subjects.

  View details for DOI 10.1371/journal.pone.0055971

  View details for Web of Science ID 000314692800061

  View details for PubMedID 23393606

  View details for PubMedCentralID PMC3564913

• Combined F-18-Fluoride and F-18-FDG PET/CT Scanning for Evaluation of Malignancy: Results of an International Multicenter Trial JOURNAL OF NUCLEAR MEDICINE Iagaru, A., Mittra, E., Mosci, C., Dick, D. W., Sathekge, M., Prakash, V., Iyer, V., Lapa, P., Isidoro, J., de Lima, J. M., Gambhir, S. S. 2013; 54 (2): 176-183

  Abstract

  (18)F-FDG PET/CT is used in a variety of cancers, but because of variable rates of glucose metabolism, not all cancers are reliably identified. (18)F(-) PET/CT allows for the acquisition of highly sensitive and specific images of the skeleton. We prospectively evaluated combined (18)F(-)/(18)F-FDG as a single PET/CT examination for evaluation of cancer patients and compared it with separate (18)F(-) PET/CT and (18)F-FDG PET/CT scans.One hundred fifteen participants with cancer were prospectively enrolled in an international multicenter trial evaluating (18)F(-) PET/CT, (18)F-FDG PET/CT, and combined (18)F(-)/(18)F-FDG PET/CT. The 3 PET/CT scans were performed sequentially within 4 wk of one another for each patient.(18)F(-)/(18)F-FDG PET/CT allowed for accurate interpretation of radiotracer uptake outside the skeleton, with findings similar to those of (18)F-FDG PET/CT. In 19 participants, skeletal disease was more extensive on (18)F(-) PET/CT and (18)F(-)/(18)F-FDG PET/CT than on (18)F-FDG PET/CT. In another 29 participants, (18)F(-) PET/CT and (18)F(-)/(18)F-FDG PET/CT showed osseous metastases where (18)F-FDG PET/CT was negative. The extent of skeletal lesions was similar in 18 participants on all 3 scans.This trial demonstrated that combined (18)F(-)/(18)F-FDG PET/CT shows promising results when compared with separate (18)F(-) PET/CT and (18)F-FDG PET/CT for evaluation of cancer patients. This result opens the possibility for improved patient care and reduction in health-care costs, as will be further evaluated in future trials.

  View details for DOI 10.2967/jnumed.112.108803

  View details for Web of Science ID 000314691200016

  View details for PubMedID 23243299

• Dissection of the role of the tumor microenvironment in oncogene addiction by ex vivo and in situ imaging AACR/SNMMI Joint Conference on State-of-the-Art Molecular Imaging in Cancer Biology and Therapy Tong, L., Jeon, J., Shen, B., Jianghong, R., Chin, F., Gambhir, S., Felsher, D. SOC NUCLEAR MEDICINE INC. 2013: 25–25

  View details for Web of Science ID 000314691400075

• Colony-stimulating factor 1 receptor (CSF1R) signaling in injured neurons facilitates protection and survival JOURNAL OF EXPERIMENTAL MEDICINE Luo, J., Elwood, F., Britschgi, M., Villeda, S., Zhang, H., Ding, Z., Zhu, L., Alabsi, H., Getachew, R., Narasimhan, R., Wabl, R., Fainberg, N., James, M. L., Wong, G., Relton, J., Gambhir, S. S., Pollard, J. W., Wyss-Coray, T. 2013; 210 (1): 157-172

  Abstract

  Colony-stimulating factor 1 (CSF1) and interleukin-34 (IL-34) are functional ligands of the CSF1 receptor (CSF1R) and thus are key regulators of the monocyte/macrophage lineage. We discovered that systemic administration of human recombinant CSF1 ameliorates memory deficits in a transgenic mouse model of Alzheimer's disease. CSF1 and IL-34 strongly reduced excitotoxin-induced neuronal cell loss and gliosis in wild-type mice when administered systemically before or up to 6 h after injury. These effects were accompanied by maintenance of cAMP responsive element-binding protein (CREB) signaling in neurons rather than in microglia. Using lineage-tracing experiments, we discovered that a small number of neurons in the hippocampus and cortex express CSF1R under physiological conditions and that kainic acid-induced excitotoxic injury results in a profound increase in neuronal receptor expression. Selective deletion of CSF1R in forebrain neurons in mice exacerbated excitotoxin-induced death and neurodegeneration. We conclude that CSF1 and IL-34 provide powerful neuroprotective and survival signals in brain injury and neurodegeneration involving CSF1R expression on neurons.

  View details for DOI 10.1084/jem.20120412

  View details for Web of Science ID 000313560900014

  View details for PubMedID 23296467

  View details for PubMedCentralID PMC3549715

• Synthesis of ligand-functionalized water-soluble [F-18]YF3 nanoparticles for PET imaging NANOSCALE Xiong, L., Shen, B., Behera, D., Gambhir, S. S., Chin, F. T., Rao, J. 2013; 5 (8): 3253-3256

  Abstract

  We report a simple, efficient synthesis of novel (18)F-labeled imaging agents based on YF3 nanoparticles. Targeting ligands and antitumor drug molecules can be introduced onto the YF3 nanoparticles in a one-pot synthesis. The (18)F-labeling reaction proceeds in aqueous solutions at room temperature with excellent radiolabeling yields (>80%) in a very short time (5-10 min). (18)F-labeled YF3 nanoparticles displayed high stability in mouse and human serum, and their application for mapping lymph nodes in live rats after local injection has also been demonstrated.

  View details for DOI 10.1039/c3nr00335c

  View details for Web of Science ID 000316959500019

  View details for PubMedID 23508229

  View details for PubMedCentralID PMC3645980

• Real-time, continuous, fluorescence sensing in a freely-moving subject with an implanted hybrid VCSEL/CMOS biosensor. Biomedical optics express O'Sullivan, T. D., Heitz, R. T., Parashurama, N., Barkin, D. B., Wooley, B. A., Gambhir, S. S., Harris, J. S., Levi, O. 2013; 4 (8): 1332-1341

  Abstract

  Performance improvements in instrumentation for optical imaging have contributed greatly to molecular imaging in living subjects. In order to advance molecular imaging in freely moving, untethered subjects, we designed a miniature vertical-cavity surface-emitting laser (VCSEL)-based biosensor measuring 1cm(3) and weighing 0.7g that accurately detects both fluorophore and tumor-targeted molecular probes in small animals. We integrated a critical enabling component, a complementary metal-oxide semiconductor (CMOS) read-out integrated circuit, which digitized the fluorescence signal to achieve autofluorescence-limited sensitivity. After surgical implantation of the lightweight sensor for two weeks, we obtained continuous and dynamic fluorophore measurements while the subject was un-anesthetized and mobile. The technology demonstrated here represents a critical step in the path toward untethered optical sensing using an integrated optoelectronic implant.

  View details for DOI 10.1364/BOE.4.001332

  View details for PubMedID 24009996

• Molecular Imaging Primer Gambhir, S. S. Apple iBookstore. 2013
• MicroRNA-Regulated Non-Viral Vectors with Improved Tumor Specifically in an Orthotopic Rat Model pf Hepatocellular Carcinoma Gene Therapy Ronald, J. A., Katzenberg, R., Nielsen, C. H., Jae, H. J., Hofmann, L. V., Gambhir, S. S. 2013: 1006-1013
• Evolution of BRET Biosensors from Live Cell to Tissue Scale Frontiers in Endocrinology De, A., Jasani, A., Arora, R., Gambhir, S. S. 2013
• Development and Validation of Non-Intergrative, Self-Limited, and Replicating Minicircles for Safe Reporter Gene Imaging of Cell-Based Therapies PLoS One Ronald, J. A., Cusso, L., Chuang, H. Y., Yan, X., Dragulescu-Andrasi, A., Gambhir, S. S. 2013: e73138
• Modification of a commercially available photoacoustic imaging system for the use of 1064nm and 532nm wavelengths to assess photoacoustic contrast agents Heinmiller, A., Homan, K., Emelianov, S., Cole, A., Gambhir, S., Needles, A., Theodoropoulos, C., Hirson, D., Oraevsky, A. A., Wang, L. V. SPIE-INT SOC OPTICAL ENGINEERING. 2013

  View details for DOI 10.1117/12.2004804

  View details for Web of Science ID 000322832800085

• Single-cell Photonic Nanocavity Probes Shambat, G., Kothapalli, R., Provine, J., Sarmiento, T., Harris, J., Gambhir, S., Vuckovic, J., IEEE IEEE. 2013

  View details for Web of Science ID 000355262503102

• Comparison of Gaussian and Poisson Noise Models in a Hybrid Reference Spectrum and Principal Component Analysis Algorithm for Raman Spectroscopy Conference on Single Molecule Spectroscopy and Superresolution Imaging VI Van de Sompel, D., Garai, E., Zavaleta, C., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2013

  View details for DOI 10.1117/12.2005455

  View details for Web of Science ID 000321741600011

• A Brain Tumor Molecular Imaging Strategy using a New Triple-Modality MRI-Photoacoustic-Raman Nanoparticle Conference on Photons Plus Ultrasound - Imaging and Sensing de la Zerda, A., Kircher, M. F., Jokerst, J. V., Zavaleta, C. L., Kempen, P. J., Mittra, E., Pitter, K., Huang, R., Campos, C., Habte, F., Sinclair, R., Brennan, C. W., Mellinghoff, I. K., Holland, E. C., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2013

  View details for DOI 10.1117/12.2001719

  View details for Web of Science ID 000322832800007

• Development and validation of non-integrative, self-limited, and replicating minicircles for safe reporter gene imaging of cell-based therapies. PloS one Ronald, J. A., Cusso, L., Chuang, H., Yan, X., Dragulescu-Andrasi, A., Gambhir, S. S. 2013; 8 (8)

  Abstract

  Reporter gene (RG) imaging of cell-based therapies provides a direct readout of therapeutic efficacy by assessing the fate of implanted cells. To permit long-term cellular imaging, RGs are traditionally required to be integrated into the cellular genome. This poses a potential safety risk and regulatory bottleneck for clinical translation as integration can lead to cellular transformation. To address this issue, we have developed non-integrative, replicating minicircles (MCs) as an alternative platform for safer monitoring of cells in living subjects. We developed both plasmids and minicircles containing the scaffold/matrix attachment regions (S/MAR) of the human interferon-beta gene, driven by the CMV promoter, and expressing the bioluminescence RG firefly luciferase. Constructs were transfected into breast cancer cells, and expanded S/MAR minicircle clones showed luciferase signal for greater than 3 months in culture and minicircles remained as episomes. Importantly, luciferase activity in clonal populations was slowly lost over time and this corresponded to a loss of episome, providing a way to reversibly label cells. To monitor cell proliferation in vivo, 1.5×10(6) cells carrying the S/MAR minicircle were implanted subcutaneously into mice (n = 5) and as tumors developed significantly more bioluminescence signal was noted at day 35 and 43 compared to day 7 post-implant (p<0.05). To our knowledge, this is the first work examining the use of episomal, self-limited, replicating minicircles to track the proliferation of cells using non-invasive imaging in living subjects. Continued development of S/MAR minicircles will provide a broadly applicable vector platform amenable with any of the numerous RG technologies available to allow therapeutic cell fate to be assessed in individual patients, and to achieve this without the need to manipulate the cell's genome so that safety concerns are minimized. This will lead to safe tools to assess treatment response at earlier time points and improve the precision of cell-based therapies.

  View details for DOI 10.1371/journal.pone.0073138

  View details for PubMedID 24015294

  View details for PubMedCentralID PMC3756008

• A transgenic tri-modality reporter mouse. PloS one Yan, X., Ray, P., Paulmurugan, R., Tong, R., Gong, Y., Sathirachinda, A., Wu, J. C., Gambhir, S. S. 2013; 8 (8)

  Abstract

  Transgenic mouse with a stably integrated reporter gene(s) can be a valuable resource for obtaining uniformly labeled stem cells, tissues, and organs for various applications. We have generated a transgenic mouse model that ubiquitously expresses a tri-fusion reporter gene (fluc2-tdTomato-ttk) driven by a constitutive chicken β-actin promoter. This "Tri-Modality Reporter Mouse" system allows one to isolate most cells from this donor mouse and image them for bioluminescent (fluc2), fluorescent (tdTomato), and positron emission tomography (PET) (ttk) modalities. Transgenic colonies with different levels of tri-fusion reporter gene expression showed a linear correlation between all three-reporter proteins (R(2)=0.89 for TdTomato vs Fluc, R(2)=0.94 for Fluc vs TTK, R(2)=0.89 for TdTomato vs TTK) in vitro from tissue lysates and in vivo by optical and PET imaging. Mesenchymal stem cells (MSCs) isolated from this transgenics showed high level of reporter gene expression, which linearly correlated with the cell numbers (R(2)=0.99 for bioluminescence imaging (BLI)). Both BLI (R(2)=0.93) and micro-PET (R(2)=0.94) imaging of the subcutaneous implants of Tri-Modality Reporter Mouse derived MSCs in nude mice showed linear correlation with the cell numbers and across different imaging modalities (R(2)=0.97). Serial imaging of MSCs transplanted to mice with acute myocardial infarction (MI) by intramyocardial injection exhibited significantly higher signals in MI heart at days 2, 3, 4, and 7 (p<0.01). MSCs transplanted to the ischemic hindlimb of nude mice showed significantly higher BLI and PET signals in the first 2 weeks that dropped by 4(th) week due to poor cell survival. However, laser Doppler perfusion imaging revealed that blood circulation in the ischemic limb was significantly improved in the MSCs transplantation group compared with the control group. In summary, this mouse can be used as a source of donor cells and organs in various research areas such as stem cell research, tissue engineering research, and organ transplantation.

  View details for DOI 10.1371/journal.pone.0073580

  View details for PubMedID 23951359

  View details for PubMedCentralID PMC3739740

• Development and application of stable phantoms for the evaluation of photoacoustic imaging instruments. PloS one Bohndiek, S. E., Bodapati, S., Van de Sompel, D., Kothapalli, S., Gambhir, S. S. 2013; 8 (9)

  Abstract

  Photoacoustic imaging combines the high contrast of optical imaging with the spatial resolution and penetration depth of ultrasound. This technique holds tremendous potential for imaging in small animals and importantly, is clinically translatable. At present, there is no accepted standard physical phantom that can be used to provide routine quality control and performance evaluation of photoacoustic imaging instruments. With the growing popularity of the technique and the advent of several commercial small animal imaging systems, it is important to develop a strategy for assessment of such instruments. Here, we developed a protocol for fabrication of physical phantoms for photoacoustic imaging from polyvinyl chloride plastisol (PVCP). Using this material, we designed and constructed a range of phantoms by tuning the optical properties of the background matrix and embedding spherical absorbing targets of the same material at different depths. We created specific designs to enable: routine quality control; the testing of robustness of photoacoustic signals as a function of background; and the evaluation of the maximum imaging depth available. Furthermore, we demonstrated that we could, for the first time, evaluate two small animal photoacoustic imaging systems with distinctly different light delivery, ultrasound imaging geometries and center frequencies, using stable physical phantoms and directly compare the results from both systems.

  View details for DOI 10.1371/journal.pone.0075533

  View details for PubMedID 24086557

  View details for PubMedCentralID PMC3783368

• Stable phantoms for characterization of photoacoustic tomography (PAT) systems Conference on Design and Performance Validation of Phantoms Used in Conjunction with Optical Measurement of Tissue V Bohndiek, S. E., Van de Sompel, D., Bodapati, S., Kothapalli, S. R., Gambhir, S. S. SPIE-INT SOC OPTICAL ENGINEERING. 2013

  View details for DOI 10.1117/12.2005195

  View details for Web of Science ID 000322903900005

• Evolution of BRET Biosensors from Live Cell to Tissue-Scale In vivo Imaging. Frontiers in endocrinology De, A., Jasani, A., Arora, R., Gambhir, S. S. 2013; 4: 131-?

  Abstract

  Development of bioluminescence resonance energy transfer (BRET) based genetic sensors for sensing biological functions such as protein-protein interactions (PPIs) in vivo has a special value in measuring such dynamic events at their native environment. Since its inception in the late nineties, BRET related research has gained significant momentum in terms of adding versatility to the assay format and wider applicability where it has been suitably used. Beyond the scope of quantitative measurement of PPIs and protein dimerization, molecular imaging applications based on BRET assays have broadened its scope for screening pharmacologically important compounds by in vivo imaging as well. In this mini-review we focus on an in-depth analysis of engineered BRET systems developed and their successful application to cell-based assays as well as in vivo non-invasive imaging in live subjects.

  View details for DOI 10.3389/fendo.2013.00131

  View details for PubMedID 24065957

• Unexpected Dissemination Patterns in Lymphoma Progression Revealed by Serial Imaging within a Murine Lymph Node CANCER RESEARCH Ito, K., Smith, B. R., Parashurama, N., Yoon, J., Song, S. Y., Miething, C., Mallick, P., Lowe, S., Gambhir, S. S. 2012; 72 (23): 6111-6118

  Abstract

  Non-Hodgkin lymphoma (NHL) is a heterogeneous and highly disseminated disease, but the mechanisms of its growth and dissemination are not well understood. Using a mouse model of this disease, we used multimodal imaging, including intravital microscopy (IVM) combined with bioluminescence, as a powerful tool to better elucidate NHL progression. We injected enhanced green fluorescent protein and luciferase-expressing Eμ-Myc/Arf(-/-) (Cdkn2a(-/-)) mouse lymphoma cells (EL-Arf(-/-)) into C57BL/6NCrl mice intravenously. Long-term observation inside a peripheral lymph node was enabled by a novel lymph node internal window chamber technique that allows chronic, sequential lymph node imaging under in vivo physiologic conditions. Interestingly, during early stages of tumor progression we found that few if any lymphoma cells homed initially to the inguinal lymph node (ILN), despite clear evidence of lymphoma cells in the bone marrow and spleen. Unexpectedly, we detected a reproducible efflux of lymphoma cells from spleen and bone marrow, concomitant with a massive and synchronous influx of lymphoma cells into the ILN, several days after injection. We confirmed a coordinated efflux/influx of tumor cells by injecting EL-Arf(-/-) lymphoma cells directly into the spleen and observing a burst of lymphoma cells, validating that the burst originated in organs remote from the lymph nodes. Our findings argue that in NHL an efflux of tumor cells from one disease site to another, distant site in which they become established occurs in discrete bursts.

  View details for DOI 10.1158/0008-5472.CAN-12-2579

  View details for Web of Science ID 000311893100005

  View details for PubMedID 23033441

  View details for PubMedCentralID PMC3664177

• Human Flexor Tendon Tissue Engineering: Revitalization of Biostatic Allograft Scaffolds TISSUE ENGINEERING PART A Woon, C. Y., Farnebo, S., Schmitt, T., Kraus, A., Megerle, K., Hung Pham, H., Yan, X., Gambhir, S. S., Chang, J. 2012; 18 (23-24): 2406-2417

  Abstract

  Cadaveric tendon allografts form a readily available and underutilized source of graft material. Because of their material properties, allografts are biomechanically and biologically superior to synthetic scaffolds. However, before clinical use, allografts must undergo decellularization to reduce immunogenicity and oxidation to increase porosity, leaving a nonvital biostatic scaffold. Ex vivo seeding, or revitalization, is thought to hasten graft incorporation and stimulate intrinsic tendon healing, permitting early mobilization and return to function. In this study, we examined physical and biochemical augmentation methods, including scaffold surface scoring (physical) and rehydration of lyophilized scaffolds in serum (biochemical). Scaffolds were divided into four groups: (1) scored scaffolds, (2) lyophilized scaffolds rehydrated in fetal calf serum (FCS), (3) scaffolds both scored and rehydrated in FCS, and (4) control scaffolds. Scaffolds were reseeded with adipose-derived stem cells (ADSCs). Reseeding efficacy was quantified by a live cell and total cell assays and qualified histologically with hematoxylin and eosin, live/dead and SYTO green nucleic acid stains, TUNEL apoptosis stains, procollagen stains, and transmission electron microscopy. Scaffold-seeded cell viability at up to 2 weeks in vitro and up to 4 weeks in vivo was demonstrated with bioluminescent imaging of scaffolds seeded with luciferase-positive ADSCs. The effect of seeding on scaffold biomechanical properties was demonstrated with evaluation of ultimate tensile stress (UTS) and an elastic modulus (EM). We found that scaffold surface scoring led to an increase in live and total cell attachment and penetration (MTS assay, p<0.001 and DNA assay, p=0.003, respectively). Histology confirmed greater total cell number in both construct core and surface in scored compared with unscored constructs. Cells reseeded on scored constructs displayed reduced apoptosis, persistent procollagen production, and had a similar ultrastructural relationship to the surrounding matrix as native tenocytes on transmission electron microscopy. Rehydration of lyophilized scaffolds in serum did not improve reseeding. Seeded constructs demonstrated greater UTS and EM than unseeded constructs. Scaffolds seeded with ADSC-luc2-eGFP demonstrated persistent viability for at least 2 weeks in vitro. In conclusion, tendon surface scoring increases surface and core reseeding in vitro and may be incorporated as a final step in allograft processing before clinical implantation.

  View details for DOI 10.1089/ten.tea.2012.0152

  View details for Web of Science ID 000311600800002

• Exogenous MC3T3 Preosteoblasts Migrate Systemically and Mitigate the Adverse Effects of Wear Particles TISSUE ENGINEERING PART A Fritton, K., Ren, P., Gibon, E., Rao, A. J., Ma, T., Biswal, S., Gambhir, S. S., Goodman, S. B. 2012; 18 (23-24): 2559-2567

  Abstract

  Understanding how relevant cell types respond to wear particles will reveal new avenues for treating osteolysis following joint replacements. In this study, we investigate the effects of ultrahigh molecular weight polyethylene (UHMWPE) particles on preosteoblast migration and function. We infused UHMWPE particles or saline into the left femur of mice and injected luciferase-expressing preosteoblasts (MC3T3 cells) into each left ventricle. Bioluminescence imaging (BLI) confirmed systemic administration of MC3T3 cells. BLI throughout the 28-day experiment showed greater MC3T3 migration to the site of particle infusion than to the site of saline infusion, with significant differences on days 0, 4, and 6 (p≤0.055). Immunostaining revealed a greater number of osteoblasts and osteoclasts in the particle-infused femora, indicating greater bone turnover. The bone mineralization of the particle-infused femora increased significantly when compared to saline-infused femora (an increase of 146.4±27.9 vs. 12.8±8.7 mg/mL, p=0.008). These results show that infused preosteoblasts can migrate to the site of wear particles. Additionally, as the migrated cells were associated with increased bone mineralization in spite of the presence of particles, increasing osteoblast recruitment is a potential strategy for combating bone loss due to increased osteoclast/macrophage number and decreased osteoblast function.

  View details for DOI 10.1089/ten.tea.2012.0086

  View details for PubMedID 22741555

• Human flexor tendon tissue engineering: revitalization of biostatic allograft scaffolds. Tissue engineering. Part A Woon, C. Y., Farnebo, S., Schmitt, T., Kraus, A., Megerle, K., Pham, H., Yan, X., Gambhir, S. S., Chang, J. 2012; 18 (23-24): 2406-2417

  Abstract

  Cadaveric tendon allografts form a readily available and underutilized source of graft material. Because of their material properties, allografts are biomechanically and biologically superior to synthetic scaffolds. However, before clinical use, allografts must undergo decellularization to reduce immunogenicity and oxidation to increase porosity, leaving a nonvital biostatic scaffold. Ex vivo seeding, or revitalization, is thought to hasten graft incorporation and stimulate intrinsic tendon healing, permitting early mobilization and return to function. In this study, we examined physical and biochemical augmentation methods, including scaffold surface scoring (physical) and rehydration of lyophilized scaffolds in serum (biochemical). Scaffolds were divided into four groups: (1) scored scaffolds, (2) lyophilized scaffolds rehydrated in fetal calf serum (FCS), (3) scaffolds both scored and rehydrated in FCS, and (4) control scaffolds. Scaffolds were reseeded with adipose-derived stem cells (ADSCs). Reseeding efficacy was quantified by a live cell and total cell assays and qualified histologically with hematoxylin and eosin, live/dead and SYTO green nucleic acid stains, TUNEL apoptosis stains, procollagen stains, and transmission electron microscopy. Scaffold-seeded cell viability at up to 2 weeks in vitro and up to 4 weeks in vivo was demonstrated with bioluminescent imaging of scaffolds seeded with luciferase-positive ADSCs. The effect of seeding on scaffold biomechanical properties was demonstrated with evaluation of ultimate tensile stress (UTS) and an elastic modulus (EM). We found that scaffold surface scoring led to an increase in live and total cell attachment and penetration (MTS assay, p<0.001 and DNA assay, p=0.003, respectively). Histology confirmed greater total cell number in both construct core and surface in scored compared with unscored constructs. Cells reseeded on scored constructs displayed reduced apoptosis, persistent procollagen production, and had a similar ultrastructural relationship to the surrounding matrix as native tenocytes on transmission electron microscopy. Rehydration of lyophilized scaffolds in serum did not improve reseeding. Seeded constructs demonstrated greater UTS and EM than unseeded constructs. Scaffolds seeded with ADSC-luc2-eGFP demonstrated persistent viability for at least 2 weeks in vitro. In conclusion, tendon surface scoring increases surface and core reseeding in vitro and may be incorporated as a final step in allograft processing before clinical implantation.

  View details for DOI 10.1089/ten.TEA.2012.0152

  View details for PubMedID 22712522

• Dimeric 18F-RGD PET Tracer for alpha v beta 3-Targeted Imaging of Experimental Abdominal Aortic Aneurysm Disease Kitagawa, T., Kosuge, H., Chang, E., James, M. L., Yamamoto, T., Gambhir, S. S., Dalman, R. L., McConnell, M. V. LIPPINCOTT WILLIAMS & WILKINS. 2012

  View details for Web of Science ID 000208885004051

• Continuous sensing of tumor-targeted molecular probes with a vertical cavity surface emitting laser-based biosensor JOURNAL OF BIOMEDICAL OPTICS Parashurama, N., O'Sullivan, T. D., de la Zerda, A., El Kalassi, P., Cho, S., Liu, H., Teed, R., Levy, H., Rosenberg, J., Cheng, Z., Levi, O., Harris, J. S., Gambhir, S. S. 2012; 17 (11)

  Abstract

  Molecular optical imaging is a widespread technique for interrogating molecular events in living subjects. However, current approaches preclude long-term, continuous measurements in awake, mobile subjects, a strategy crucial in several medical conditions. Consequently, we designed a novel, lightweight miniature biosensor for in vivo continuous optical sensing. The biosensor contains an enclosed vertical-cavity surface-emitting semiconductor laser and an adjacent pair of near-infrared optically filtered detectors. We employed two sensors (dual sensing) to simultaneously interrogate normal and diseased tumor sites. Having established the sensors are precise with phantom and in vivo studies, we performed dual, continuous sensing in tumor (human glioblastoma cells) bearing mice using the targeted molecular probe cRGD-Cy5.5, which targets αVβ3 cell surface integrins in both tumor neovasculature and tumor. The sensors capture the dynamic time-activity curve of the targeted molecular probe. The average tumor to background ratio after signal calibration for cRGD-Cy5.5 injection is approximately 2.43±0.95 at 1 h and 3.64±1.38 at 2 h (N=5 mice), consistent with data obtained with a cooled charge coupled device camera. We conclude that our novel, portable, precise biosensor can be used to evaluate both kinetics and steady state levels of molecular probes in various disease applications.

  View details for DOI 10.1117/1.JBO.17.11.117004

  View details for Web of Science ID 000314502700046

  View details for PubMedID 23123976

  View details for PubMedCentralID PMC3595658

• Gold Nanorods for Ovarian Cancer Detection with Photoacoustic Imaging and Resection Guidance via Raman Imaging in Living Mice ACS NANO Jokerst, J. V., Cole, A. J., Van de Sompel, D., Gambhir, S. S. 2012; 6 (11): 10366-10377

  Abstract

  Improved imaging approaches are needed for ovarian cancer screening, diagnosis, staging, and resection guidance. Here, we propose a combined photoacoustic (PA)/Raman approach using gold nanorods (GNRs) as a passively targeted molecular imaging agent. GNRs with three different aspect ratios were studied. Those with an aspect ratio of 3.5 were selected for their highest ex vivo and in vivo PA signal and used to image subcutaneous xenografts of the 2008, HEY, and SKOV3 ovarian cancer cell lines in living mice. Maximum PA signal was observed within 3 h for all three lines tested and increased signal persisted for at least two days postadministration. There was a linear relationship (R(2) = 0.95) between the PA signal and the concentration of injected molecular imaging agent with a calculated limit of detection of 0.40 nM GNRs in the 2008 cell line. The same molecular imaging agent could be used for clear visualization of the margin between tumor and normal tissue and tumor debulking via surface-enhanced Raman spectroscopy (SERS) imaging. Finally, we validated the imaging findings with biodistribution data and elemental analysis. To the best of our knowledge, this is the first report of in vivo imaging of ovarian cancer tumors with a photoacoustic and Raman imaging agent.

  View details for DOI 10.1021/nn304347g

  View details for Web of Science ID 000311521700112

  View details for PubMedID 23101432

  View details for PubMedCentralID PMC3572720

• New Positron Emission Tomography (PET) Radioligand for Imaging sigma-1 Receptors in Living Subjects JOURNAL OF MEDICINAL CHEMISTRY James, M. L., Shen, B., Zavaleta, C. L., Nielsen, C. H., Mesangeau, C., Vuppala, P. K., Chan, C., Avery, B. A., Fishback, J. A., Matsumoto, R. R., Gambhir, S. S., McCurdy, C. R., Chin, F. T. 2012; 55 (19): 8272-8282

  Abstract

  σ-1 receptor (S1R) radioligands have the potential to detect and monitor various neurological diseases. Herein we report the synthesis, radiofluorination, and evaluation of a new S1R ligand 6-(3-fluoropropyl)-3-(2-(azepan-1-yl)ethyl)benzo[d]thiazol-2(3H)-one ([(18)F]FTC-146, [(18)F]13). [(18)F]13 was synthesized by nucleophilic fluorination, affording a product with >99% radiochemical purity (RCP) and specific activity (SA) of 2.6 ± 1.2 Ci/μmol (n = 13) at end of synthesis (EOS). Positron emission tomography (PET) and ex vivo autoradiography studies of [(18)F]13 in mice showed high uptake of the radioligand in S1R rich regions of the brain. Pretreatment with 1 mg/kg haloperidol (2), nonradioactive 13, or BD1047 (18) reduced the binding of [(18)F]13 in the brain at 60 min by 80%, 82%, and 81%, respectively, suggesting that [(18)F]13 accumulation in mouse brain represents specific binding to S1Rs. These results indicate that [(18)F]13 is a promising candidate radiotracer for further evaluation as a tool for studying S1Rs in living subjects.

  View details for DOI 10.1021/jm300371c

  View details for Web of Science ID 000309643500008

  View details for PubMedID 22853801

• Improving Image Quality by Accounting for Changes in Water Temperature during a Photoacoustic Tomography Scan PLOS ONE Van de Sompel, D., Sasportas, L. S., Dragulescu-Andrasi, A., Bohndiek, S., Gambhir, S. S. 2012; 7 (10)

  Abstract

  The emerging field of photoacoustic tomography is rapidly evolving with many new system designs and reconstruction algorithms being published. Many systems use water as a coupling medium between the scanned object and the ultrasound transducers. Prior to a scan, the water is heated to body temperature to enable small animal imaging. During the scan, the water heating system of some systems is switched off to minimize the risk of bubble formation, which leads to a gradual decrease in water temperature and hence the speed of sound. In this work, we use a commercially available scanner that follows this procedure, and show that a failure to model intra-scan temperature decreases as small as 1.5°C leads to image artifacts that may be difficult to distinguish from true structures, particularly in complex scenes. We then improve image quality by continuously monitoring the water temperature during the scan and applying variable speed of sound corrections in the image reconstruction algorithm. While upgrading to an air bubble-free heating pump and keeping it running during the scan could also solve the changing temperature problem, we show that a software correction for the temperature changes provides a cost-effective alternative to a hardware upgrade. The efficacy of the software corrections was shown to be consistent across objects of widely varying appearances, namely physical phantoms, ex vivo tissue, and in vivo mouse imaging. To the best of our knowledge, this is the first study to demonstrate the efficacy of modeling temporal variations in the speed of sound during photoacoustic scans, as opposed to spatial variations as focused on by previous studies. Since air bubbles pose a common problem in ultrasonic and photoacoustic imaging systems, our results will be useful to future small animal imaging studies that use scanners with similarly limited heating units.

  View details for DOI 10.1371/journal.pone.0045337

  View details for Web of Science ID 000309807700008

  View details for PubMedID 23071512

  View details for PubMedCentralID PMC3469660

• The Impact of Partial Volume Correction in the Evaluation of Solitary Pulmonary Nodules by FDG PET/CT in a Population at Intermediate Risk of Lung Cancer 4th International Symposium on Targeted Radiotherapy and Dosimetry (ISTARD) in Conjunction with the 25th Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Keu, K., Nair, V. S., Mittra, E., Gambhir, S. S., Iagaru, A. SPRINGER. 2012: S455–S455

  View details for Web of Science ID 000309726602293

• Feasibility of Limited Thoracic FDG PET/CT for the Evaluation of Solitary Pulmonary Nodules in Patients With Intermediate and High Risk of Lung Cancer Keu, K., Mittra, E., Gambhir, S. S., Iagaru, A. SPRINGER. 2012: S455

  View details for Web of Science ID 000309726602292

• FEASIBILITY OF AN INTRAMOLECULAR COMPLEMENTATION STRATEGY FOR SPLIT-REPORTER GENE IMAGING OF DRUGGABLE PROTEIN MISFOLDING IN BRAIN CANCER 17th Annual Scientific Meeting and Education Day of the Society-for-Neuro-Oncology (SNO) Massoud, T. F., Paulmurugan, R., Gambhir, S. S. OXFORD UNIV PRESS INC. 2012: 11–11

  View details for Web of Science ID 000310971300048

• Exploratory Clinical Trial of (4S)-4-(3-[F-18]fluoropropyl)-L-glutamate for Imaging x(C) Transporter Using Positron Emission Tomography in Patients with Non-Small Cell Lung or Breast Cancer CLINICAL CANCER RESEARCH Baek, S., Choi, C., Ahn, S. H., Lee, J. W., Gong, G., Ryu, J., Oh, S. J., Bacher-Stier, C., Fels, L., Koglin, N., Hultsch, C., Schatz, C. A., Dinkelborg, L. M., Mittra, E. S., Gambhir, S. S., Moon, D. H. 2012; 18 (19): 5427-5437

  Abstract

  (4S)-4-(3-[(18)F]fluoropropyl)-l-glutamate (BAY 94-9392, alias [(18)F]FSPG) is a new tracer to image x(C)(-) transporter activity with positron emission tomography (PET). We aimed to explore the tumor detection rate of [(18)F]FSPG in patients relative to 2-[(18)F]fluoro-2-deoxyglucose ([(18)F]FDG). The correlation of [(18)F]FSPG uptake with immunohistochemical expression of x(C)(-) transporter and CD44, which stabilizes the xCT subunit of system x(C)(-), was also analyzed.Patients with non-small cell lung cancer (NSCLC, n = 10) or breast cancer (n = 5) who had a positive [(18)F]FDG uptake were included in this exploratory study. PET images were acquired following injection of approximately 300 MBq [(18)F]FSPG. Immunohistochemistry was done using xCT- and CD44-specific antibody.[(18)F]FSPG PET showed high uptake in the kidney and pancreas with rapid blood clearance. [(18)F]FSPG identified all 10 NSCLC and three of the five breast cancer lesions that were confirmed by pathology. [(18)F]FSPG detected 59 of 67 (88%) [(18)F]FDG lesions in NSCLC, and 30 of 73 (41%) in breast cancer. Seven lesions were additionally detected only on [(18)F]FSPG in NSCLC. The tumor-to-blood pool standardized uptake value (SUV) ratio was not significantly different from that of [(18)F]FDG in NSCLC; however, in breast cancer, it was significantly lower (P < 0.05). The maximum SUV of [(18)F]FSPG correlated significantly with the intensity of immunohistochemical staining of x(C)(-) transporter and CD44 (P < 0.01).[(18)F]FSPG seems to be a promising tracer with a relatively high cancer detection rate in patients with NSCLC. [(18)F]FSPG PET may assess x(C)(-) transporter activity in patients with cancer.

  View details for DOI 10.1158/1078-0432.CCR-12-0214

  View details for Web of Science ID 000311906600027

  View details for PubMedID 22893629

• Intraoperative Imaging of Tumors Using Cerenkov Luminescence Endoscopy: A Feasibility Experimental Study JOURNAL OF NUCLEAR MEDICINE Liu, H., Carpenter, C. M., Jiang, H., Pratx, G., Sun, C., Buchin, M. P., Gambhir, S. S., Xing, L., Cheng, Z. 2012; 53 (10): 1579-1584

  Abstract

  Cerenkov luminescence imaging (CLI) is an emerging new molecular imaging modality that is relatively inexpensive, easy to use, and has high throughput. CLI can image clinically available PET and SPECT probes using optical instrumentation. Cerenkov luminescence endoscopy (CLE) is one of the most intriguing applications that promise potential clinical translation. We developed a prototype customized fiberscopic Cerenkov imaging system to investigate the potential in guiding minimally invasive surgical resection.All experiments were performed in a dark chamber. Cerenkov luminescence from (18)F-FDG samples containing decaying radioactivity was transmitted through an optical fiber bundle and imaged by an intensified charge-coupled device camera. Phantoms filled with (18)F-FDG were used to assess the imaging spatial resolution. Finally, mice bearing subcutaneous C6 glioma cells were injected intravenously with (18)F-FDG to determine the feasibility of in vivo imaging. The tumor tissues were exposed, and CLI was performed on the mouse before and after surgical removal of the tumor using the fiber-based imaging system and compared with a commercial optical imaging system.The sensitivity of this particular setup was approximately 45 kBq (1.21 μCi)/300 μL. The 3 smallest sets of cylindric holes in a commercial SPECT phantom were identifiable via this system, demonstrating that the system has a resolution better than 1.2 mm. Finally, the in vivo tumor imaging study demonstrated the feasibility of using CLI to guide the resection of tumor tissues.This proof-of-concept study explored the feasibility of using fiber-based CLE for the detection of tumor tissue in vivo for guided surgery. With further improvements of the imaging sensitivity and spatial resolution of the current system, CLE may have a significant application in the clinical setting in the near future.

  View details for DOI 10.2967/jnumed.111.098541

  View details for Web of Science ID 000309432400017

  View details for PubMedID 22904353

• Positron Emission Tomography of Cu-64-DOTA-Rituximab in a Transgenic Mouse Model Expressing Human CD20 for Clinical Translation to Image NHL MOLECULAR IMAGING AND BIOLOGY Natarajan, A., Gowrishankar, G., Nielsen, C. H., Wang, S., Iagaru, A., Goris, M. L., Gambhir, S. S. 2012; 14 (5): 608-616

  Abstract

  This study aims to evaluate (64)Cu-DOTA-rituximab (PETRIT) in a preclinical transgenic mouse model expressing human CD20 for potential clinical translation.(64)Cu was chelated to DOTA-rituximab. Multiple radiolabeling, quality assurance, and imaging experiments were performed. The human CD20 antigen was expressed in B cells of transgenic mice (CD20TM). The mice groups studied were: (a) control (nude mice, n = 3) that received 7.4 MBq/dose, (b) with pre-dose (CD20TM, n = 6) received 2 mg/kg pre-dose of cold rituximab prior to PETRIT of 7.4 MBq/dose, and (c) without pre-dose (CD20TM, n = 6) PETRIT alone received 7.4 MBq/dose. Small animal PET was used to image mice at various time points (0, 1, 2, 4, 24, 48, and 72 h). The OLINDA/EXM software was used to determine the human equivalent dose for individual organs.PETRIT was obtained with a specific activity of 545 ± 38.91 MBq/nmole, radiochemical purity >95%, and immunoreactivity >75%. At 24 h, spleenic uptake of PETRIT%ID/g (mean ± STD) with and without pre-dose was 1.76 ± 0.43% and 16.5 ± 0.45%, respectively (P value = 0.01). Liver uptake with and without pre-dose was 0.41 ± 0.51% and 0.52 ± 0.17% (P value = 0.86), respectively. The human equivalents of highest dose organs with and without pre-dose are osteogenic cells at 30.8 ± 0.4 μSv/MBq and the spleen at 99 ± 4 μSv/MBq, respectively.PET imaging with PETRIT in huCD20 transgenic mice provided human dosimetry data for eventual applications in non-Hodgkins lymphoma patients.

  View details for DOI 10.1007/s11307-011-0537-8

  View details for Web of Science ID 000308819300011

  View details for PubMedID 22231277

• alpha v beta 3 Integrins as a Biomarker of Disease Recurrence in Glioblastoma Multiforme: Initial Clinical Results Using 18F FPPRGD2 PET/CT 4th International Symposium on Targeted Radiotherapy and Dosimetry (ISTARD) in Conjunction with the 25th Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Iagaru, A., Mosci, C., Mittra, E. S., Shin, B., Chin, F., Gambhir, S. S. SPRINGER. 2012: S244–S245

  View details for Web of Science ID 000309726600324

• Remodeling of Endogenous Mammary Epithelium by Breast Cancer Stem Cells STEM CELLS Parashurama, N., Lobo, N. A., Ito, K., Mosley, A. R., Habte, F. G., Zabala, M., Smith, B. R., Lam, J., Weissman, I. L., Clarke, M. F., Gambhir, S. S. 2012; 30 (10): 2114-2127

  Abstract

  Poorly regulated tissue remodeling results in increased breast cancer risk, yet how breast cancer stem cells (CSC) participate in remodeling is unknown. We performed in vivo imaging of changes in fluorescent, endogenous duct architecture as a metric for remodeling. First, we quantitatively imaged physiologic remodeling of primary branches of the developing and regenerating mammary tree. To assess CSC-specific remodeling events, we isolated CSC from MMTV-Wnt1 (mouse mammary tumor virus long-term repeat enhancer driving Wnt1 oncogene) breast tumors, a well studied model in which tissue remodeling affects tumorigenesis. We confirm that CSC drive tumorigenesis, suggesting a link between CSC and remodeling. We find that normal, regenerating, and developing gland maintain a specific branching pattern. In contrast, transplantation of CSC results in changes in the branching patterns of endogenous ducts while non-CSC do not. Specifically, in the presence of CSC, we identified an increased number of branches, branch points, ducts which have greater than 40 branches (5/33 for CSC and 0/39 for non-CSC), and histological evidence of increased branching. Moreover, we demonstrate that only CSC implants invade into surrounding stroma with structures similar to developing mammary ducts (nine for CSC and one for non-CSC). Overall, we demonstrate a novel approach for imaging physiologic and pathological remodeling. Furthermore, we identify unique, CSC-specific, remodeling events. Our data suggest that CSC interact with the microenvironment differently than non-CSC, and that this could eventually be a therapeutic approach for targeting CSC.

  View details for DOI 10.1002/stem.1205

  View details for Web of Science ID 000308928300005

  View details for PubMedID 22899386

• Designed hydrophilic and charge mutations of the fibronectin domain: towards tailored protein biodistribution PROTEIN ENGINEERING DESIGN & SELECTION Hackel, B. J., Sathirachinda, A., Gambhir, S. S. 2012; 25 (10): 639-647

  Abstract

  Engineered proteins are attractive affinity scaffolds for molecular imaging and drug delivery. Although exquisite binding specificity and affinity can be engineered, many proteins exhibit off-target uptake, particularly in the kidneys and liver, from physiologic effects. We quantified the ability to alter renal and hepatic uptake via hydrophilic and charge mutations. As a model protein, we used the 10th type III domain of human fibronectin, which has been engineered to bind many targets and has been validated for molecular imaging. We screened rational mutants, identified by structural and phylogenetic analyses, to yield eight mutations that collectively substantially increase protein hydrophilicity. Mutation of two parental clones yielded four domains with a range of hydrophilicity. These proteins were labeled with (64)Cu, injected intravenously into nu/nu mice (n = 3-5 each) and evaluated by positron emission tomography. Renal uptake strongly correlated with hydrophilicity (Pearson's correlation coefficient = 0.97), ranging from 29 ± 11 to 100 ± 22% ID/g at 1 h. Hepatic uptake inversely correlated with hydrophilicity (Pearson's correlation coefficient = -0.92), ranging from 30 ± 7 to 3 ± 1% ID/g. Thus, renal and hepatic uptake are directly tunable through hydrophilic mutation, identifiable by structural and phylogenetic analyses. To investigate charge, we mutated acidic and basic residues in both parental clones and evaluated (64)Cu-labeled mutants in nu/nu mice (n = 5-7). Selected charge removal reduced kidney signal: 78 ± 13 to 51 ± 8%ID/g (P < 0.0001) for the hydrophilic clone and 32 ± 10 to 21 ± 3 (P = 0.0005) for the hydrophobic clone. Elucidation of hydrophilicity and charge enabled modulation of background signal thereby enhancing the utility of protein scaffolds as translatable targeting agents for molecular imaging and therapy.

  View details for DOI 10.1093/protein/gzs036

  View details for Web of Science ID 000309468100016

  View details for PubMedID 22691700

  View details for PubMedCentralID PMC3449399

• Microfluidic Single-Cell Analysis Shows That Porcine Induced Pluripotent Stem Cell-Derived Endothelial Cells Improve Myocardial Function by Paracrine Activation CIRCULATION RESEARCH Gu, M., Nguyen, P. K., Lee, A. S., Xu, D., Hu, S., Plews, J. R., Han, L., Huber, B. C., Lee, W. H., Gong, Y., de Almeida, P. E., Lyons, J., Ikeno, F., Pacharinsak, C., Connolly, A. J., Gambhir, S. S., Robbins, R. C., Longaker, M. T., Wu, J. C. 2012; 111 (7): 882-893

  Abstract

  Induced pluripotent stem cells (iPSCs) hold great promise for the development of patient-specific therapies for cardiovascular disease. However, clinical translation will require preclinical optimization and validation of large-animal iPSC models.To successfully derive endothelial cells from porcine iPSCs and demonstrate their potential utility for the treatment of myocardial ischemia.Porcine adipose stromal cells were reprogrammed to generate porcine iPSCs (piPSCs). Immunohistochemistry, quantitative PCR, microarray hybridization, and angiogenic assays confirmed that piPSC-derived endothelial cells (piPSC-ECs) shared similar morphological and functional properties as endothelial cells isolated from the autologous pig aorta. To demonstrate their therapeutic potential, piPSC-ECs were transplanted into mice with myocardial infarction. Compared with control, animals transplanted with piPSC-ECs showed significant functional improvement measured by echocardiography (fractional shortening at week 4: 27.2±1.3% versus 22.3±1.1%; P<0.001) and MRI (ejection fraction at week 4: 45.8±1.3% versus 42.3±0.9%; P<0.05). Quantitative protein assays and microfluidic single-cell PCR profiling showed that piPSC-ECs released proangiogenic and antiapoptotic factors in the ischemic microenvironment, which promoted neovascularization and cardiomyocyte survival, respectively. Release of paracrine factors varied significantly among subpopulations of transplanted cells, suggesting that transplantation of specific cell populations may result in greater functional recovery.In summary, this is the first study to successfully differentiate piPSCs-ECs from piPSCs and demonstrate that transplantation of piPSC-ECs improved cardiac function after myocardial infarction via paracrine activation. Further development of these large animal iPSC models will yield significant insights into their therapeutic potential and accelerate the clinical translation of autologous iPSC-based therapy.

  View details for DOI 10.1161/CIRCRESAHA.112.269001

  View details for PubMedID 22821929

• Discovery and validation of small-molecule heat-shock protein 90 inhibitors through multimodality molecular imaging in living subjects PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Chan, C. T., Reeves, R. E., Geller, R., Yaghoubi, S. S., Hoehne, A., Solow-Cordero, D. E., Chiosis, G., Massoud, T. F., Paulmurugan, R., Gambhir, S. S. 2012; 109 (37): E2476-E2485

  Abstract

  Up-regulation of the folding machinery of the heat-shock protein 90 (Hsp90) chaperone protein is crucial for cancer progression. The two Hsp90 isoforms (α and β) play different roles in response to chemotherapy. To identify isoform-selective inhibitors of Hsp90(α/β)/cochaperone p23 interactions, we developed a dual-luciferase (Renilla and Firefly) reporter system for high-throughput screening (HTS) and monitoring the efficacy of Hsp90 inhibitors in cell culture and live mice. HTS of a 30,176 small-molecule chemical library in cell culture identified a compound, N-(5-methylisoxazol-3-yl)-2-[4-(thiophen-2-yl)-6-(trifluoromethyl)pyrimidin-2-ylthio]acetamide (CP9), that binds to Hsp90(α/β) and displays characteristics of Hsp90 inhibitors, i.e., degradation of Hsp90 client proteins and inhibition of cell proliferation, glucose metabolism, and thymidine kinase activity, in multiple cancer cell lines. The efficacy of CP9 in disrupting Hsp90(α/β)/p23 interactions and cell proliferation in tumor xenografts was evaluated by non-invasive, repetitive Renilla luciferase and Firefly luciferase imaging, respectively. At 38 h posttreatment (80 mg/kg × 3, i.p.), CP9 led to selective disruption of Hsp90α/p23 as compared with Hsp90β/p23 interactions. Small-animal PET/CT in the same cohort of mice showed that CP9 treatment (43 h) led to a 40% decrease in (18)F-fluorodeoxyglucose uptake in tumors relative to carrier control-treated mice. However, CP9 did not lead to significant degradation of Hsp90 client proteins in tumors. We performed a structural activity relationship study with 62 analogs of CP9 and identified A17 as the lead compound that outperformed CP9 in inhibiting Hsp90(α/β)/p23 interactions in cell culture. Our efforts demonstrated the power of coupling of HTS with multimodality molecular imaging and led to identification of Hsp90 inhibitors.

  View details for DOI 10.1073/pnas.1205459109

  View details for PubMedID 22895790

• Cationic versus Neutral Microbubbles for Ultrasound-mediated Gene Delivery in Cancer RADIOLOGY Wang, D. S., Panje, C., Pysz, M. A., Paulmurugan, R., Rosenberg, J., Gambhir, S. S., Schneider, M., Willmann, J. K. 2012; 264 (3): 721-732

  Abstract

  To test whether plasmid-binding cationic microbubbles (MBs) enhance ultrasound-mediated gene delivery efficiency relative to control neutral MBs in cell culture and in vivo tumors in mice.Animal studies were approved by the institutional animal care committee. Cationic and neutral MBs were characterized in terms of size, charge, circulation time, and DNA binding. Click beetle luciferase (CBLuc) reporter plasmids were mixed with cationic or neutral MBs. The ability of cationic MBs to protect bound plasmids from nuclease degradation was tested by means of a deoxyribonuclease (DNase) protection assay. Relative efficiencies of ultrasound-mediated transfection (ultrasound parameters: 1 MHz, 1 W/cm(2), 20% duty cycle, 1 minute) of CBLuc to endothelial cells by using cationic, neutral, or no MBs were compared in cell culture. Ultrasound-mediated gene delivery to mouse hind limb tumors was performed in vivo (n = 24) with insonation (1 MHz, 2 W/cm(2), 50% duty cycle, 5 minutes) after intravenous administration of CBLuc with cationic, neutral, or no MBs. Tumor luciferase activity was assessed by means of serial in vivo bioluminescence imaging and ex vivo analysis. Results were compared by using analysis of variance.Cationic MBs (+15.8 mV; DNA binding capacity, 0.03 pg per MB) partially protected bound DNA from DNase degradation. Mean CBLuc expression of treated endothelial cells in culture was 20-fold higher with cationic than with neutral MBs (219.0 relative light units [RLUs]/µg protein ± 92.5 [standard deviation] vs 10.9 RLUs/µg protein ± 2.7, P = .001) and was significantly higher (P < .001) than that in the no MB and no ultrasound control groups. Serial in vivo bioluminescence of mouse tumors was significantly higher with cationic than with neutral MBs ([5.9 ± 2.2] to [9.3 ± 5.2] vs [2.4 ± 0.8] to [2.9 ± 1.1] × 10(4) photons/sec/cm(2)/steradian, P < .0001) and versus no MB and no ultrasound controls (P < .0001). Results of ex vivo analysis confirmed these results (ρ = 0.88, P < .0001).Plasmid-binding cationic MBs enhance ultrasound-mediated gene delivery efficiency relative to neutral MBs in both cell culture and mouse hind limb tumors.

  View details for DOI 10.1148/radiol.12112368

  View details for Web of Science ID 000308645500013

  View details for PubMedID 22723497

  View details for PubMedCentralID PMC3426857

• Circulating tumour cells in early breast cancer LANCET ONCOLOGY Nair, V. S., Keu, K. V., Kuhn, P., Gambhir, S. S. 2012; 13 (9): E370-E371

  View details for Web of Science ID 000308425600024

  View details for PubMedID 22935234

• Tissue-engineered collateral ligament composite allografts for scapholunate ligament reconstruction: an experimental study. journal of hand surgery Endress, R., Woon, C. Y., Farnebo, S. J., Behn, A., Bronstein, J., Pham, H., Yan, X., Gambhir, S. S., Chang, J. 2012; 37 (8): 1529-1537

  Abstract

  In patients with chronic scapholunate (SL) dissociation or dynamic instability, ligament repair is often not possible, and surgical reconstruction is indicated. The ideal graft ligament would recreate both anatomical and biomechanical properties of the dorsal scapholunate ligament (dorsal SLIL). The finger proximal interphalangeal joint (PIP joint) collateral ligament could possibly be a substitute ligament.We harvested human PIP joint collateral ligaments and SL ligaments from 15 cadaveric limbs. We recorded ligament length, width, and thickness, and measured the biomechanical properties (ultimate load, stiffness, and displacement to failure) of native dorsal SLIL, untreated collateral ligaments, decellularized collateral ligaments, and SL repairs with bone-collateral ligament-bone composite collateral ligament grafts. As proof of concept, we then reseeded decellularized bone-collateral ligament-bone composite grafts with green fluorescent protein-labeled adipo-derived mesenchymal stem cells and evaluated them histologically.There was no difference in ultimate load, stiffness, and displacement to failure among native dorsal SLIL, untreated and decellularized collateral ligaments, and SL repairs with tissue-engineered collateral ligament grafts. With pair-matched untreated and decellularized scaffolds, there was no difference in ultimate load or stiffness. However, decellularized ligaments revealed lower displacement to failure compared with untreated ligaments. There was no difference in displacement between decellularized ligaments and native dorsal SLIL. We successfully decellularized grafts with recently described techniques, and they could be similarly reseeded.Proximal interphalangeal joint collateral ligament-based bone-collateral ligament-bone composite allografts had biomechanical properties similar to those of native dorsal SLIL. Decellularization did not adversely affect material properties.These tissue-engineered grafts may offer surgeons another option for reconstruction of chronic SL instability.

  View details for DOI 10.1016/j.jhsa.2012.05.020

  View details for PubMedID 22835583

• Tissue-engineered Collateral Ligament Composite Allografts for Scapholunate Ligament Reconstruction: An Experimental Study JOURNAL OF HAND SURGERY-AMERICAN VOLUME Endress, R., Woon, C. Y., Farnebo, S. J., Behn, A., Bronstein, J., Pham, H., Yan, X., Gambhir, S. S., Chang, J. 2012; 37A (8): 1529-1537

  Abstract

  In patients with chronic scapholunate (SL) dissociation or dynamic instability, ligament repair is often not possible, and surgical reconstruction is indicated. The ideal graft ligament would recreate both anatomical and biomechanical properties of the dorsal scapholunate ligament (dorsal SLIL). The finger proximal interphalangeal joint (PIP joint) collateral ligament could possibly be a substitute ligament.We harvested human PIP joint collateral ligaments and SL ligaments from 15 cadaveric limbs. We recorded ligament length, width, and thickness, and measured the biomechanical properties (ultimate load, stiffness, and displacement to failure) of native dorsal SLIL, untreated collateral ligaments, decellularized collateral ligaments, and SL repairs with bone-collateral ligament-bone composite collateral ligament grafts. As proof of concept, we then reseeded decellularized bone-collateral ligament-bone composite grafts with green fluorescent protein-labeled adipo-derived mesenchymal stem cells and evaluated them histologically.There was no difference in ultimate load, stiffness, and displacement to failure among native dorsal SLIL, untreated and decellularized collateral ligaments, and SL repairs with tissue-engineered collateral ligament grafts. With pair-matched untreated and decellularized scaffolds, there was no difference in ultimate load or stiffness. However, decellularized ligaments revealed lower displacement to failure compared with untreated ligaments. There was no difference in displacement between decellularized ligaments and native dorsal SLIL. We successfully decellularized grafts with recently described techniques, and they could be similarly reseeded.Proximal interphalangeal joint collateral ligament-based bone-collateral ligament-bone composite allografts had biomechanical properties similar to those of native dorsal SLIL. Decellularization did not adversely affect material properties.These tissue-engineered grafts may offer surgeons another option for reconstruction of chronic SL instability.

  View details for DOI 10.1016/j.jhsa.2012.05.020

  View details for Web of Science ID 000307260200001

• Impact of Screening Test Performance and Cost on Mortality Reduction and Cost-effectiveness of Multimodal Ovarian Cancer Screening CANCER PREVENTION RESEARCH Drescher, C. W., Hawley, S., Thorpe, J. D., Marticke, S., McIntosh, M., Gambhir, S. S., Urban, N. 2012; 5 (8): 1015-1024

  Abstract

  Ongoing ovarian cancer screening trials are investigating the efficacy of a two-step screening strategy using currently available blood and imaging tests [CA125 and transvaginal sonography (TVS)]. Concurrently, efforts to develop new biomarkers and imaging tests seek to improve screening performance beyond its current limits. This study estimates the mortality reduction, years of life saved, and cost-effectiveness achievable by annual multimodal screening using increasing CA125 to select women for TVS, and predicts improvements achievable by replacing currently available screening tests with hypothetical counterparts with better performance characteristics. An existing stochastic microsimulation model is refined and used to screen a virtual cohort of 1 million women from ages 45 to 85 years. Each woman is assigned a detailed disease course and screening results timeline. The preclinical behavior of CA125 and TVS is simulated using empirical data derived from clinical trials. Simulations in which the disease incidence and performance characteristics of the screening tests are independently varied are conducted to evaluate the impact of these factors on overall screening performance and costs. Our results show that when applied to women at average risk, annual screening using increasing CA125 to select women for TVS achieves modest mortality reduction (~13%) and meets currently accepted cost-effectiveness guidelines. Screening outcomes are relatively insensitive to second-line test performance and costs. Identification of a first-line test that does substantially better than CA125 and has similar costs is required for screening to reduce ovarian mortality by at least 25% and be reasonably cost-effective.

  View details for DOI 10.1158/1940-6207.CAPR-11-0468

  View details for Web of Science ID 000308223500004

  View details for PubMedID 22750949

• Fluorescent Magnetic Nanoparticles for Magnetically Enhanced Cancer Imaging and Targeting in Living Subjects ACS NANO Fu, A., Wilson, R. J., Smith, B. R., Mullenix, J., Earhart, C., Akin, D., Guccione, S., Wang, S. X., Gambhir, S. S. 2012; 6 (8): 6862-6869

  Abstract

  Early detection and targeted therapy are two major challenges in the battle against cancer. Novel imaging contrast agents and targeting approaches are greatly needed to improve the sensitivity and specificity of cancer theranostic agents. Here, we implemented a novel approach using a magnetic micromesh and biocompatible fluorescent magnetic nanoparticles (FMN) to magnetically enhance cancer targeting in living subjects. This approach enables magnetic targeting of systemically administered individual FMN, containing a single 8 nm superparamagnetic iron oxide core. Using a human glioblastoma mouse model, we show that nanoparticles can be magnetically retained in both the tumor neovasculature and surrounding tumor tissues. Magnetic accumulation of nanoparticles within the neovasculature was observable by fluorescence intravital microscopy in real time. Finally, we demonstrate that such magnetically enhanced cancer targeting augments the biological functions of molecules linked to the nanoparticle surface.

  View details for DOI 10.1021/nn301670a

  View details for Web of Science ID 000307988900039

  View details for PubMedID 22857784

  View details for PubMedCentralID PMC3601027

• Shape Matters: Intravital Microscopy Reveals Surprising Geometrical Dependence for Nanoparticles in Tumor Models of Extravasation NANO LETTERS Smith, B. R., Kempen, P., Bouley, D., Xu, A., Liu, Z., Melosh, N., Dai, H., Sinclair, R., Gambhir, S. S. 2012; 12 (7): 3369-3377

  Abstract

  Delivery is one of the most critical obstacles confronting nanoparticle use in cancer diagnosis and therapy. For most oncological applications, nanoparticles must extravasate in order to reach tumor cells and perform their designated task. However, little understanding exists regarding the effect of nanoparticle shape on extravasation. Herein we use real-time intravital microscopic imaging to meticulously examine how two different nanoparticles behave across three different murine tumor models. The study quantitatively demonstrates that high-aspect ratio single-walled carbon nanotubes (SWNTs) display extravasational behavior surprisingly different from, and counterintuitive to, spherical nanoparticles although the nanoparticles have similar surface coatings, area, and charge. This work quantitatively indicates that nanoscale extravasational competence is highly dependent on nanoparticle geometry and is heterogeneous.

  View details for DOI 10.1021/nl204175t

  View details for Web of Science ID 000306296200004

  View details for PubMedID 22650417

  View details for PubMedCentralID PMC3495189

• In vivo targeting of HER2-positive tumor using 2-helix affibody molecules AMINO ACIDS Ren, G., Webster, J. M., Liu, Z., Zhang, R., Miao, Z., Liu, H., Gambhir, S. S., Syud, F. A., Cheng, Z. 2012; 43 (1): 405-413

  Abstract

  Molecular imaging of human epidermal growth factor receptor type 2 (HER2) expression has drawn significant attention because of the unique role of the HER2 gene in diagnosis, therapy and prognosis of human breast cancer. In our previous research, a novel cyclic 2-helix small protein, MUT-DS, was discovered as an anti-HER2 Affibody analog with high affinity through rational protein design and engineering. MUT-DS was then evaluated for positron emission tomography (PET) of HER2-positive tumor by labeling with two radionuclides, 68Ga and 18F, with relatively short half-life (t1/2<2 h). In order to fully study the in vivo behavior of 2-helix small protein and demonstrate that it could be a robust platform for labeling with a variety of radionuclides for different applications, in this study, MUT-DS was further radiolabeled with 64Cu or 111In and evaluated for in vivo targeting of HER2-positive tumor in mice. Design 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) conjugated MUT-DS (DOTA-MUT-DS) was chemically synthesized using solid phase peptide synthesizer and I2 oxidation. DOTA-MUT-DS was then radiolabeled with 64Cu or 111In to prepare the HER2 imaging probe (64Cu/111In-DOTA-MUT-DS). Both biodistribution and microPET imaging of the probe were evaluated in nude mice bearing subcutaneous HER2-positive SKOV3 tumors. DOTA-MUT-DS could be successfully synthesized and radiolabeled with 64Cu or 111In. Biodistribution study showed that tumor uptake value of 64Cu or 111In-labeled DOTA-MUT-DS was 4.66±0.38 or 2.17±0.15%ID/g, respectively, in nude mice bearing SKOV3 xenografts (n=3) at 1 h post-injection (p.i.). Tumor-to-blood and tumor-to-muscle ratios for 64Cu-DOTA-MUT-DS were attained to be 3.05 and 3.48 at 1 h p.i., respectively, while for 111In-DOTA-MUT-DS, they were 2.04 and 3.19, respectively. Co-injection of the cold Affibody molecule ZHER2:342 with 64Cu-DOTA-MUT-DS specifically reduced the SKOV3 tumor uptake of the probe by 48%. 111In-DOTA-MUT-DS displayed lower liver uptake at all the time points investigated and higher tumor to blood ratios at 4 and 20 h p.i., when compared with 64Cu-DOTA-MUT-DS. This study demonstrates that the 2-helix protein based probes, 64Cu/111In DOTA-MUT-DS, are promising molecular probes for imaging HER2-positive tumor. Two-helix small protein scaffold holds great promise as a novel and robust platform for imaging and therapy applications.

  View details for DOI 10.1007/s00726-011-1096-7

  View details for Web of Science ID 000305210800041

  View details for PubMedID 21984380

• Photoacoustic Imaging of Mesenchymal Stem Cells in Living Mice via Silica-Coated Gold Nanorods ACS NANO Jokerst, J. V., Thangaraj, M., Kempen, P. J., Sinclair, R., Gambhir, S. S. 2012; 6 (7): 5920-5930

  Abstract

  Improved imaging modalities are critically needed for optimizing stem cell therapy. Techniques with real-time content to guide and quantitate cell implantation are especially important in applications such as musculoskeletal regenerative medicine. Here, we report the use of silica-coated gold nanorods as a contrast agent for photoacoustic imaging and quantitation of mesenchymal stem cells in rodent muscle tissue. The silica coating increased the uptake of gold into the cell more than 5-fold, yet no toxicity or proliferation changes were observed in cells loaded with this contrast agent. Pluripotency of the cells was retained, and secretome analysis indicated that only IL-6 was disregulated more than 2-fold from a pool of 26 cytokines. The low background of the technique allowed imaging of down to 100,000 cells in vivo. The spatial resolution is 340 μm, and the temporal resolution is 0.2 s, which is at least an order of magnitude below existing cell imaging approaches. This approach has significant advantages over traditional cell imaging techniques like positron emission tomography and magnetic resonance imaging including real time monitoring of stem cell therapy.

  View details for DOI 10.1021/nn302042y

  View details for Web of Science ID 000306673800020

  View details for PubMedID 22681633

  View details for PubMedCentralID PMC3582222

• A Hybrid Least Squares and Principal Component Analysis Algorithm for Raman Spectroscopy PLOS ONE Van de Sompel, D., Garai, E., Zavaleta, C., Gambhir, S. S. 2012; 7 (6)

  Abstract

  Raman spectroscopy is a powerful technique for detecting and quantifying analytes in chemical mixtures. A critical part of Raman spectroscopy is the use of a computer algorithm to analyze the measured Raman spectra. The most commonly used algorithm is the classical least squares method, which is popular due to its speed and ease of implementation. However, it is sensitive to inaccuracies or variations in the reference spectra of the analytes (compounds of interest) and the background. Many algorithms, primarily multivariate calibration methods, have been proposed that increase robustness to such variations. In this study, we propose a novel method that improves robustness even further by explicitly modeling variations in both the background and analyte signals. More specifically, it extends the classical least squares model by allowing the declared reference spectra to vary in accordance with the principal components obtained from training sets of spectra measured in prior characterization experiments. The amount of variation allowed is constrained by the eigenvalues of this principal component analysis. We compare the novel algorithm to the least squares method with a low-order polynomial residual model, as well as a state-of-the-art hybrid linear analysis method. The latter is a multivariate calibration method designed specifically to improve robustness to background variability in cases where training spectra of the background, as well as the mean spectrum of the analyte, are available. We demonstrate the novel algorithm's superior performance by comparing quantitative error metrics generated by each method. The experiments consider both simulated data and experimental data acquired from in vitro solutions of Raman-enhanced gold-silica nanoparticles.

  View details for DOI 10.1371/journal.pone.0038850

  View details for Web of Science ID 000305583300060

  View details for PubMedID 22723895

  View details for PubMedCentralID PMC3377733

• Development of a Novel Long-Lived ImmunoPET Tracer for Monitoring Lymphoma Therapy in a Humanized Transgenic Mouse Model. Bioconjugate chemistry Natarajan, A., Habte, F., Gambhir, S. S. 2012

  Abstract

  Positron emission tomography (PET) is an attractive imaging tool to localize and quantify tracer biodistribution. ImmunoPET with an intact mAb typically requires two to four days to achieve optimized tumor-to-normal ratios. Thus, a positron emitter with a half-life of two to four days such as zirconium-89 [(89)Zr] (t(1/2): 78.4 h) is ideal. We have developed an antibody-based, long-lived immunoPET tracer (89)Zr-Desferrioxamine-p-SCN (Df-Bz-NCS)-rituximab (Zr-iPET) to image tumor for longer durations in a humanized CD20-expressing transgenic mouse model. To optimize the radiolabeling efficiency of (89)Zr with Df-Bz-rituximab, multiple radiolabelings were performed. Radiochemical yield, purity, immunoreactivity, and stability assays were carried out to characterize the Zr-iPET for chemical and biological integrity. This tracer was used to image transgenic mice that express the human CD20 on their B cells (huCD20TM). Each huCD20TM mouse received a 7.4 MBq/dose. One group (n = 3) received a 2 mg/kg predose (blocking) of cold rituximab 2 h prior to (89)Zr-iPET; the other group (n = 3) had no predose (nonblocking). Small animal PET/CT was used to image mice at 1, 4, 24, 48, 72, and 120 h. Quality assurance of the (89)Zr-iPET demonstrated NCS-Bz-Df: antibody ratio (c/a: 1.5 ± 0.31), specific activity (0.44-1.64 TBq/mol), radiochemical yield (>70%), and purity (>98%). The Zr-iPET immunoreactivity was >80%. At 120 h, Zr-iPET uptake (% ID/g) as mean ± STD for blocking and nonblocking groups in spleen was 3.2 ± 0.1% and 83.3 ± 2.0% (p value <0.0013.). Liver uptake was 1.32 ± 0.05% and 0.61 ± 0.001% (p value <0.0128) for blocking and nonblocking, respectively. The small animal PET/CT image shows the spleen specific uptake of Zr-iPET in mice at 120 h after tracer injection. Compared to the liver, the spleen specific uptake of Zr-iPET is very high due to the expression of huCD20. We optimized the radiolabeling efficiency of (89)Zr with Df-Bz-rituximab. These radioimmunoconjugate lots were stable up to 5 days in serum in vitro. The present study showed that (89)Zr is well-suited for mAbs to image cancer over an extended period of time (up to 5 days).

  View details for DOI 10.1021/bc300039r

  View details for PubMedID 22621257

  View details for PubMedCentralID PMC3459285

• Endoscopic imaging of Cerenkov luminescence BIOMEDICAL OPTICS EXPRESS Kothapalli, S., Liu, H., Liao, J. C., Cheng, Z., Gambhir, S. S. 2012; 3 (6): 1215-1225

  Abstract

  We demonstrate feasibility of endoscopic imaging of Cerenkov light originated when charged nuclear particles, emitted from radionuclides, travel through a biological tissue of living subjects at superluminal velocity. The endoscopy imaging system consists of conventional optical fiber bundle/ clinical endoscopes, an optical imaging lens system, and a sensitive low-noise charge coupled device (CCD) camera. Our systematic studies using phantom samples show that Cerenkov light from as low as 1 µCi of radioactivity emitted from (18)F-Fluorodeoxyglucose (FDG) can be coupled and transmitted through conventional optical fibers and endoscopes. In vivo imaging experiments with tumor bearing mice, intravenously administered with (18)F-FDG, further demonstrated that Cerenkov luminescence endoscopy is a promising new tool in the field of endoscopic molecular imaging.

  View details for Web of Science ID 000304965700007

  View details for PubMedID 22741069

  View details for PubMedCentralID PMC3370963

• Transatlantic Consensus Group on active surveillance and focal therapy for prostate cancer BJU INTERNATIONAL Ahmed, H. U., Akin, O., Coleman, J. A., Crane, S., Emberton, M., Goldenberg, L., Hricak, H., Kattan, M. W., Kurhanewicz, J., Moore, C. M., Parker, C., Polascik, T. J., Scardino, P., Van As, N., Villers, A. 2012; 109 (11): 1636-1647

  Abstract

  What's known on the subject? and What does the study add? Active surveillance for prostate cancer is gaining increasing acceptance for low risk prostate cancer. Focal therapy is an emerging tissue preservation strategy that aims for treat only areas of cancer. Early phase trials have shown that side-effects can be significantly reduced using focal therapy. There is significant uncertainty in both active surveillance and focal therapy. This consensus group paper provides a road-map for clinical practice and research for both tissue-preserving strategies in the areas of patient population, tools for risk stratification and cancer localisation, treatment interventions as well as comparators and outcome measures in future comparative trials.To reach consensus on key issues for clinical practice and future research in active surveillance and focal therapy in managing localized prostate cancer.A group of expert urologists, oncologists, radiologists, pathologists and computer scientists from North America and Europe met to discuss issues in patient population, interventions, comparators and outcome measures to use in both tissue-preserving strategies of active surveillance and focal therapy. Break-out sessions were formed to provide agreement or highlight areas of disagreement on individual topics which were then collated by a writing group into statements that formed the basis of this report and agreed upon by the whole Transatlantic Consensus Group.The Transatlantic group propose that emerging diagnostic tools such as precision imaging and transperineal prostate mapping biopsy can improve prostate cancer care. These tools should be integrated into prostate cancer management and research so that better risk stratification and more effective treatment allocation can be applied. The group envisaged a process of care in which active surveillance, focal therapy, and radical treatments lie on a continuum of complementary therapies for men with a range of disease grades and burdens, rather than being applied in the mutually exclusive and competitive way they are now.The changing landscape of prostate cancer epidemiology requires the medical community to re-evaluate the entire prostate cancer diagnostic and treatment pathway in order to minimize harms resulting from over-diagnosis and over-treatment. Precise risk stratification at every point in this pathway is required alongside paradigm shifts in our thinking about what constitutes cancer in the prostate.

  View details for DOI 10.1111/j.1464-410X.2011.10633.x

  View details for Web of Science ID 000303598400015

  View details for PubMedID 22077593

• Family of Enhanced Photoacoustic Imaging Agents for High-Sensitivity and Multiplexing Studies in Living Mice ACS NANO de la Zerda, A., Bodapati, S., Teed, R., May, S. Y., Tabakman, S. M., Liu, Z., Khuri-Yakub, B. T., Chen, X., Dai, H., Gambhir, S. S. 2012; 6 (6): 4694-4701

  Abstract

  Photoacoustic imaging is a unique modality that overcomes to a great extent the resolution and depth limitations of optical imaging while maintaining relatively high contrast. However, since many diseases will not manifest an endogenous photoacoustic contrast, it is essential to develop exogenous photoacoustic contrast agents that can target diseased tissue(s). Here we present a family of novel photoacoustic contrast agents that are based on the binding of small optical dyes to single-walled carbon nanotubes (SWNT-dye). We synthesized five different SWNT-dye contrast agents using different optical dyes, creating five "flavors" of SWNT-dye nanoparticles. In particular, SWNTs that were coated with either QSY(21) (SWNT-QSY) or indocyanine green (SWNT-ICG) exhibited over 100-times higher photoacoustic contrast in living animals compared to plain SWNTs, leading to subnanomolar sensitivities. We then conjugated the SWNT-dye conjugates with cyclic Arg-Gly-Asp peptides to molecularly target the α(v)β(3) integrin, which is associated with tumor angiogenesis. Intravenous administration of these tumor-targeted imaging agents to tumor-bearing mice showed significantly higher photoacoustic signal in the tumor than in mice injected with the untargeted contrast agent. Finally, we were able to spectrally separate the photoacoustic signals of SWNT-QSY and SWNT-ICG in living animals injected subcutaneously with both particles in the same location, opening the possibility for multiplexing in vivo studies.

  View details for DOI 10.1021/nn204352r

  View details for Web of Science ID 000305661300017

  View details for PubMedID 22607191

  View details for PubMedCentralID PMC3397693

• A photonic crystal cavity-optical fiber tip nanoparticle sensor for biomedical applications APPLIED PHYSICS LETTERS Shambat, G., Kothapalli, S. R., Khurana, A., Provine, J., Sarmiento, T., Cheng, K., Cheng, Z., Harris, J., Daldrup-Link, H., Gambhir, S. S., Vuckovic, J. 2012; 100 (21)

  View details for DOI 10.1063/1.4719520

  View details for Web of Science ID 000304489900085

• Effect of a CCR1 receptor antagonist on systemic trafficking of MSCs and polyethylene particle-associated bone loss BIOMATERIALS Gibon, E., Yao, Z., Rao, A. J., Zwingenberger, S., Batke, B., Valladares, R., Smith, R. L., Biswal, S., Gambhir, S. S., Goodman, S. B. 2012; 33 (14): 3632-3638

  Abstract

  Particle-associated periprosthetic osteolysis remains a major issue in joint replacement. Ongoing bone loss resulting from wear particle-induced inflammation is accompanied by continued attempts at bone repair. Previously we showed that mesenchymal stem cells (MSCs) are recruited systemically to bone exposed to continuous infusion of ultra high molecular weight polyethylene (UHMWPE) particles. The chemokine-receptor axis that mediates this process is unknown. We tested two hypotheses: (1) the CCR1 receptor mediates the systemic recruitment of MSCs to UHMWPE particles and (2) recruited MSCs are able to differentiate into functional mature osteoblasts and decrease particle-associated bone loss. Nude mice were allocated randomly to four groups. UHMWPE particles were continuously infused into the femoral shaft using a micro-pump. Genetically modified murine wild type reporter MSCs were injected systemically via the left ventricle. Non-invasive imaging was used to assay MSC migration and bone mineral density. Bioluminescence and immunohistochemistry confirmed the chemotaxis of reporter cells and their differentiation into mature osteoblasts in the presence of infused particles. Injection of a CCR1 antagonist decreased reporter cell recruitment to the UHMWPE particle infusion site and increased osteolysis. CCR1 appears to be a critical receptor for chemotaxis of MSCs in the presence of UHMWPE particles. Interference with CCR1 exacerbates particle-induced bone loss.

  View details for DOI 10.1016/j.biomaterials.2012.02.003

  View details for PubMedID 22364730

• 18F FPPRGD2 in GBM: Imaging alpha v beta 3 integrin levels as a biomarker of disease recurrence Iagaru, A., Mosci, C., Mittra, E., Shen, B., Chin, F., Fischbein, N., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680204100

• [18F]FPRGD2 PET/CT imaging of integrin alpha v beta 3 in renal carcinomas: Correlation with histopathology Withofs, N., Signolle, N., Nzaramba, E., Thonon, D., Leonard, M., Aerts, J., Waltregny, D., Cataldo, D., Gambhir, S., Hustinx, R. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680203206

• 18F FDG PET/CT in the management of patients with post-transplant lymphoproliferative disorder Takehana, C., Mittra, E., Quon, A., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680201186

• Evaluation of the 18F L-glutamate derivative 18F-FSPG (BAY 94-9392) in lymphoma and colon cancer patients Kumar, M., Mosci, C., Keu, K., Iagaru, A., Koglin, N., Fels, L., Bacher-Stier, C., Chin, F., Gambhir, S., Mittra, E. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680201366

• Preliminary results of [18F]FPRGD2 PET/CT imaging of integrin alpha v beta 3 levels in patients with locally advanced rectal carcinoma Withofs, N., Martinive, P., Scagnol, I., Thonon, D., Giacomelli, F., Mievis, F., Coucke, P., Cataldo, D., Gambhir, S., Hustinx, R. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680203262

• Characterization of physiological 18F-FSPG uptake in healthy volunteers: Kinetics and biodistribution Mosei, C., Kumar, M., Koglin, N., Fels, L., Bacher-Stier, C., Smolarz, K., Schwaiger, M., Gambhir, S., Mittra, E. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680201182

• Evaluation of NaF PET/CT, FDG PET/CT, combined NaF/FDG PET/CT and CT alone for detection of bone metastases Sampath, S., Sampath, S., Lutz, A., Willmann, J., Mittra, E., Gambhir, S., Iagaru, A. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680201213

• Evaluation of the 18F-labeled L-glutamate derivative 18F-FSPG (BAY 94-9392) in brain and head and neck cancer patients Kumar, M., Mosci, C., Keu, K., Iagaru, A., Koglin, N., Fels, L., Bacher-Stier, C., Chin, F., Gambhir, S., Mittra, E. SOC NUCLEAR MEDICINE INC. 2012

  View details for Web of Science ID 000443680200262

• Intratumoral versus Intravenous Gene Therapy Using a Transcriptionally Targeted Viral Vector in an Orthotopic Hepatocellular Carcinoma Rat Model JOURNAL OF VASCULAR AND INTERVENTIONAL RADIOLOGY Kim, Y. I., Ahn, B., Ronald, J. A., Katzenberg, R., Singh, A., Paulmurugan, R., Ray, S., Gambhir, S. S., Hofmann, L. V. 2012; 23 (5): 704-711

  Abstract

  To evaluate the feasibility of intratumoral delivery of adenoviral vector carrying a bidirectional two-step transcriptional amplification (TSTA) system to amplify transcriptional strength of cancer-specific Survivin promoter in a hepatocellular carcinoma model.MCA-RH7777 cells were implanted in rat liver, and tumor formation was confirmed with [(18)F]fluorodeoxyglucose (18F-FDG) positron emission tomography (PET). The adenoviral vector studied had Survivin promoter driving a therapeutic gene (tumor necrosis factor-α-related apoptosis-inducing ligand [TRAIL]) and a reporter gene (firefly luciferase [FL]; Ad-pSurvivin-TSTA-TRAIL-FL). Tumor-bearing rats were administered Ad-pSurvivin-TSTA-TRAIL-FL intravenously (n = 7) or intratumorally (n = 8). For control groups, adenovirus FL under cytomegalovirus (CMV) promoter (Ad-pCMV-FL) was administered intravenously (n = 3) or intratumorally (n = 3). One day after delivery, bioluminescence imaging was performed to evaluate transduction. At 4 and 7 days after delivery, 18F-FDG-PET was performed to evaluate therapeutic efficacy.With intravenous delivery, Ad-pSurvivin-TSTA-TRAIL-FL showed no measurable liver tumor FL signal on day 1 after delivery, but showed better therapeutic efficacy than Ad-pCMV-FL on day 7 (PET tumor/liver ratio, 3.5 ± 0.58 vs 6.0 ± 0.71; P = .02). With intratumoral delivery, Ad-pSurvivin-TSTA-TRAIL-FL showed positive FL signal from all tumors and better therapeutic efficacy than Ad-pCMV-FL on day 7 (2.4 ± 0.50 vs 5.4 ± 0.78; P = .01). In addition, intratumoral delivery of Ad-pSurvivin-TSTA-TRAIL-FL demonstrated significant decrease in tumoral viability compared with intravenous delivery (2.4 ± 0.50 vs 3.5 ± 0.58; P = .03).Intratumoral delivery of a transcriptionally targeted therapeutic vector for amplifying tumor-specific effect demonstrated better transduction efficiency and therapeutic efficacy for liver cancer than systemic delivery, and may lead to improved therapeutic outcome for future clinical practice.

  View details for DOI 10.1016/j.jvir.2012.01.053

  View details for Web of Science ID 000303557000020

  View details for PubMedID 22387029

  View details for PubMedCentralID PMC4132166

• A brain tumor molecular imaging strategy using a new triple-modality MRI-photoacoustic-Raman nanoparticle NATURE MEDICINE Kircher, M. F., de la Zerda, A., Jokerst, J. V., Zavaleta, C. L., Kempen, P. J., Mittra, E., Pitter, K., Huang, R., Campos, C., Habte, F., Sinclair, R., Brennan, C. W., Mellinghoff, I. K., Holland, E. C., Gambhir, S. S. 2012; 18 (5): 829-U235

  Abstract

  The difficulty in delineating brain tumor margins is a major obstacle in the path toward better outcomes for patients with brain tumors. Current imaging methods are often limited by inadequate sensitivity, specificity and spatial resolution. Here we show that a unique triple-modality magnetic resonance imaging-photoacoustic imaging-Raman imaging nanoparticle (termed here MPR nanoparticle) can accurately help delineate the margins of brain tumors in living mice both preoperatively and intraoperatively. The MPRs were detected by all three modalities with at least a picomolar sensitivity both in vitro and in living mice. Intravenous injection of MPRs into glioblastoma-bearing mice led to MPR accumulation and retention by the tumors, with no MPR accumulation in the surrounding healthy tissue, allowing for a noninvasive tumor delineation using all three modalities through the intact skull. Raman imaging allowed for guidance of intraoperative tumor resection, and a histological correlation validated that Raman imaging was accurately delineating the brain tumor margins. This new triple-modality-nanoparticle approach has promise for enabling more accurate brain tumor imaging and resection.

  View details for DOI 10.1038/nm.2721

  View details for Web of Science ID 000303763500053

  View details for PubMedID 22504484

  View details for PubMedCentralID PMC3422133

• Deep Tissue Photoacoustic Imaging Using a Miniaturized 2-D Capacitive Micromachined Ultrasonic Transducer Array IEEE TRANSACTIONS ON BIOMEDICAL ENGINEERING Kothapalli, S., Ma, T., Vaithilingam, S., Oralkan, O., Khuri-Yakub, B. T., Gambhir, S. S. 2012; 59 (5): 1199-1204

  Abstract

  In this paper, we demonstrate 3-D photoacoustic imaging (PAI) of light absorbing objects embedded as deep as 5 cm inside strong optically scattering phantoms using a miniaturized (4 mm × 4 mm × 500 μm), 2-D capacitive micromachined ultrasonic transducer (CMUT) array of 16 × 16 elements with a center frequency of 5.5 MHz. Two-dimensional tomographic images and 3-D volumetric images of the objects placed at different depths are presented. In addition, we studied the sensitivity of CMUT-based PAI to the concentration of indocyanine green dye at 5 cm depth inside the phantom. Under optimized experimental conditions, the objects at 5 cm depth can be imaged with SNR of about 35 dB and a spatial resolution of approximately 500 μm. Results demonstrate that CMUTs with integrated front-end amplifier circuits are an attractive choice for achieving relatively high depth sensitivity for PAI.

  View details for DOI 10.1109/TBME.2012.2183593

  View details for Web of Science ID 000303201000001

  View details for PubMedID 22249594

• Twist1 Suppresses Senescence Programs and Thereby Accelerates and Maintains Mutant Kras-Induced Lung Tumorigenesis PLOS GENETICS Tran, P. T., Shroff, E. H., Burns, T. F., Thiyagarajan, S., Das, S. T., Zabuawala, T., Chen, J., Cho, Y., Luong, R., Tamayo, P., Salih, T., Aziz, K., Adam, S. J., Vicent, S., Nielsen, C. H., Withofs, N., Sweet-Cordero, A., Gambhir, S. S., Rudin, C. M., Felsher, D. W. 2012; 8 (5)

  Abstract

  KRAS mutant lung cancers are generally refractory to chemotherapy as well targeted agents. To date, the identification of drugs to therapeutically inhibit K-RAS have been unsuccessful, suggesting that other approaches are required. We demonstrate in both a novel transgenic mutant Kras lung cancer mouse model and in human lung tumors that the inhibition of Twist1 restores a senescence program inducing the loss of a neoplastic phenotype. The Twist1 gene encodes for a transcription factor that is essential during embryogenesis. Twist1 has been suggested to play an important role during tumor progression. However, there is no in vivo evidence that Twist1 plays a role in autochthonous tumorigenesis. Through two novel transgenic mouse models, we show that Twist1 cooperates with Kras(G12D) to markedly accelerate lung tumorigenesis by abrogating cellular senescence programs and promoting the progression from benign adenomas to adenocarcinomas. Moreover, the suppression of Twist1 to physiological levels is sufficient to cause Kras mutant lung tumors to undergo senescence and lose their neoplastic features. Finally, we analyzed more than 500 human tumors to demonstrate that TWIST1 is frequently overexpressed in primary human lung tumors. The suppression of TWIST1 in human lung cancer cells also induced cellular senescence. Hence, TWIST1 is a critical regulator of cellular senescence programs, and the suppression of TWIST1 in human tumors may be an effective example of pro-senescence therapy.

  View details for DOI 10.1371/journal.pgen.1002650

  View details for Web of Science ID 000304864000004

  View details for PubMedID 22654667

  View details for PubMedCentralID PMC3360067

• MC3T3-E1 Osteoprogenitor Cells Systemically Migrate to a Bone Defect and Enhance Bone Healing TISSUE ENGINEERING PART A Gibon, E., Batke, B., Jawad, M. U., Fritton, K., Rao, A., Yao, Z., Biswal, S., Gambhir, S. S., Goodman, S. B. 2012; 18 (9-10): 968-973

  Abstract

  Although iliac crest autologous bone graft remains the gold standard for treatment of bone defects, delayed- and nonunions, and arthrodeses, several alternative strategies have been attempted, including the use of mesenchymal stem cells. Whether cells from the osteoblast lineage demonstrate systemic recruitment to an acute bone defect or fracture, and whether these cells directly participate in bone healing is controversial. This study tests two hypotheses: (1) that exogenous murine MC3T3-E1 osteoprogenitor cells with a high propensity for osteoblast differentiation are able to systemically migrate to a bone defect and (2) that the migrated MC3T3-E1 cells enhance bone healing. Two groups of nude mice were used; a bone defect was drilled in the left femoral shaft in both groups. MC3T3-E1 were used as reporter cells and injected in the left ventricle of the heart, to avoid sequestration in the lungs. Injection of saline served as a control. We used bioluminescence and microCT to assay cell recruitment and bone mineral density (BMD). Immunohistochemical staining was used to confirm the migration of reporter cells. MC3T3-E1 cells were found to systemically migrate to the bone defect. Further, BMD at the defect was significantly increased when cells were injected. Systemic cell therapy using osteoprogenitor cells may be a potential strategy to enhance bone healing.

  View details for DOI 10.1089/ten.tea.2011.0545

  View details for PubMedID 22129134

• Glioblastoma Therapy with Cytotoxic Mesenchymal Stromal Cells Optimized by Bioluminescence Imaging of Tumor and Therapeutic Cell Response PLOS ONE Alieva, M., Bago, J. R., Aguilar, E., Soler-Botija, C., Vila, O. F., Molet, J., Gambhir, S. S., Rubio, N., Blanco, J. 2012; 7 (4)

  Abstract

  Genetically modified adipose tissue derived mesenchymal stromal cells (hAMSCs) with tumor homing capacity have been proposed for localized therapy of chemo- and radiotherapy resistant glioblastomas. We demonstrate an effective procedure to optimize glioblastoma therapy based on the use of genetically modified hAMSCs and in vivo non invasive monitoring of tumor and therapeutic cells. Glioblastoma U87 cells expressing Photinus pyralis luciferase (Pluc) were implanted in combination with hAMSCs expressing a trifunctional Renilla reniformis luciferase-red fluorescent protein-thymidine kinase reporter in the brains of SCID mice that were subsequently treated with ganciclovir (GCV). The resulting optimized therapy was effective and monitoring of tumor cells by bioluminescence imaging (BLI) showed that after 49 days GCV treatment reduced significantly the hAMSC treated tumors; by a factor of 10(4) relative to controls. Using a Pluc reporter regulated by an endothelial specific promoter and in vivo BLI to image hAMSC differentiation we gained insight on the therapeutic mechanism. Implanted hAMSCs homed to tumor vessels, where they differentiated to endothelial cells. We propose that the tumor killing efficiency of genetically modified hAMSCs results from their association with the tumor vascular system and should be useful vehicles to deliver localized therapy to glioblastoma surgical borders following tumor resection.

  View details for DOI 10.1371/journal.pone.0035148

  View details for Web of Science ID 000305347400033

  View details for PubMedID 22529983

  View details for PubMedCentralID PMC3328467

• Pilot clinical trials of FSPG (BAY 94-9392): An 18F-labeled glutamate derivative for PET imaging of system xCactivity in tumors Baek, S., Mittra, E., Cho, C., Gong, G., Oh, S., Mosci, C., Kumar, M., Chin, F. T., Fels, L. M., Stephens, A. W., Koglin, N., Mueller, A., Dinkelborg, L. M., Moon, D., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2012

  View details for Web of Science ID 000209701600355

• Pilot clinical trials of FSPG (BAY 94-9392): An 18F-labeled glutamate derivative for PET imaging of system xC-activity in tumors Baek, S., Mittra, E., Choi, C., Gong, G., Oh, S., Mosci, C., Kumar, M., Chin, F. T., Fels, L. M., Stephens, A. W., Koglin, N., Mueller, A., Dinkelborg, L. M., Moon, D., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2012

  View details for DOI 10.1158/1538-7445.AM2012-LB-227

  View details for Web of Science ID 000209701601440

• A novel rotational Raman imaging device for early cancer detection Garai, E., Zavaleta, C., Sensarn, S., Mandella, M., Gambhir, S. S., Contag, C. H. AMER ASSOC CANCER RESEARCH. 2012

  View details for DOI 10.1158/1538-7445.AM2012-LB-512

  View details for Web of Science ID 000209701504209

• Early detection of pancreatic cancer in transgenic mice with ultrasonic molecular imaging and VEGFR2-targeted microbubbles Pysz, M. A., Seeley, E. S., Foygel, K., Gambhir, S. S., Tian, L., Brentnall, T. A., Willmann, J. K. AMER ASSOC CANCER RESEARCH. 2012

  View details for DOI 10.1158/1538-7445.AM2012-LB-509

  View details for Web of Science ID 000209701504206

• Correlating circulating tumor cells with F-18-FDG positron emission tomography (PET) uptake in patients with treatment naive non-small cell lung cancer: A pilot study Kuhn, P., Keu, K., Nair, V. S., Luttgen, M., Maestas, S., Bethel, K., Souder, K., Vasanawala, M., Kuschner, W., Iagaru, A. H., Hoh, C., Nieva, J., Bazhenova, L., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2012

  View details for DOI 10.1158/1538-7445.AM2012-2369

  View details for Web of Science ID 000209701505199

• A MOLECULAR IMAGING PRIMER: MODALITIES, IMAGING AGENTS, AND APPLICATIONS PHYSIOLOGICAL REVIEWS James, M. L., Gambhir, S. S. 2012; 92 (2): 897-965

  Abstract

  Molecular imaging is revolutionizing the way we study the inner workings of the human body, diagnose diseases, approach drug design, and assess therapies. The field as a whole is making possible the visualization of complex biochemical processes involved in normal physiology and disease states, in real time, in living cells, tissues, and intact subjects. In this review, we focus specifically on molecular imaging of intact living subjects. We provide a basic primer for those who are new to molecular imaging, and a resource for those involved in the field. We begin by describing classical molecular imaging techniques together with their key strengths and limitations, after which we introduce some of the latest emerging imaging modalities. We provide an overview of the main classes of molecular imaging agents (i.e., small molecules, peptides, aptamers, engineered proteins, and nanoparticles) and cite examples of how molecular imaging is being applied in oncology, neuroscience, cardiology, gene therapy, cell tracking, and theranostics (therapy combined with diagnostics). A step-by-step guide to answering biological and/or clinical questions using the tools of molecular imaging is also provided. We conclude by discussing the grand challenges of the field, its future directions, and enormous potential for further impacting how we approach research and medicine.

  View details for DOI 10.1152/physrev.00049.2010

  View details for Web of Science ID 000306562500009

  View details for PubMedID 22535898

• Use of Cu-64-labeled Fibronectin Domain with EGFR-Overexpressing Tumor Xenograft: Molecular Imaging RADIOLOGY Hackel, B. J., Kimura, R. H., Gambhir, S. S. 2012; 263 (1): 179-188

  Abstract

  To assess the ability of an engineered epidermal growth factor receptor (EGFR)-binding fibronectin domain to serve as a positron emission tomographic (PET) probe for molecular imaging of EGFR in a xenograft mouse model.An EGFR-binding fibronectin domain (fibronectin abbreviated to Fn when bound) was site-specifically labeled with copper 64 ((64)Cu) (8 MBq/nmol). Copper 64-Fn binding was tested in cell cultures with varying EGFR expression. Stability in human and mouse serum was measured in vitro. Animal experiments were approved by the Stanford University Institutional Animal Care and Use Committee. Copper 64-Fn (approximately 2 MBq) was used for PET in mice (n = 5) bearing EGFR-overexpressing xenografted tumors (approximately 5-10 mm in diameter). Results of tomography were compared with those of ex vivo gamma counting of dissected tissues. Statistical analysis was performed with t tests and adjustment for multiple comparisons.Copper 64-Fn exhibited EGFR-dependent binding to multiple cell lines in culture. The tracer was stable for 24 hours in human and mouse serum at 37°C. The tracer exhibited good tumor localization (3.4% injected dose [ID]/g ± 1.0 [standard deviation] at 1 hour), retention (2.7% ID/g ± 0.6 at 24 hours), and specificity (8.6 ± 3.0 tumor-to-muscle ratio, 8.9 ± 4.7 tumor-to-blood ratio at 1 hour). Specific targeting was verified with low localization to low-expressing MDA-MB-435 tumors (0.7% ID/g ± 0.8 at 1 hour, P = .018); specificity was further demonstrated, as a nonbinding control fibronectin had low localization to EGFR-overexpressing xenografts (0.8% ID/g ± 0.2 at 1 hour, P = .013).The stability, low background, and target-specific tumor uptake and retention of the engineered fibronectin domain make it a promising EGFR molecular imaging agent. More broadly, it validates the fibronectin domain as a potential scaffold for a generation of various molecular imaging agents.

  View details for DOI 10.1148/radiol.12111504

  View details for Web of Science ID 000302642700018

  View details for PubMedID 22344401

  View details for PubMedCentralID PMC3309798

• Prospective Evaluation of Tc-99m MDP Scintigraphy, F-18 NaF PET/CT, and F-18 FDG PET/CT for Detection of Skeletal Metastases MOLECULAR IMAGING AND BIOLOGY Iagaru, A., Mittra, E., Dick, D. W., Gambhir, S. S. 2012; 14 (2): 252-259

  Abstract

  Technetium (Tc) methylene diphosphonate (MDP) has been the standard method for bone scintigraphy for three decades. (18)F sodium fluoride ((18)F NaF) positron emission tomography (PET)/computed tomography (CT) has better resolution and is considered superior. The role of 2-deoxy-2-[(18)F]fluoro-D-glucose ((18)F FDG) PET/CT is proven in a variety of cancers, for which it has changed the practice of oncology. There are few prospective studies comparing these three methods of detection of skeletal metastases. Thus, we were prompted to initiate this prospective pilot trial.This is a prospective study (Sep 2007-Dec 2010) of 52 patients with proven malignancy referred for evaluation of skeletal metastases. There were 37 men and 15 women, 19-84 years old (average, 55.6 ± 15.9). Technetium-99m ((99m)Tc) MDP bone scintigraphy, (18)F NaF PET/CT, and (18)F FDG PET/CT were subsequently performed within 1 month.Skeletal lesions were detected by (99m)Tc MDP bone scintigraphy in 22 of 52 patients, by (18)F NaF PET/CT in 24 of 52 patients, and by (18)F FDG PET/CT in 16 of 52 patients. The image quality and evaluation of extent of disease were superior by (18)F NaF PET/CT over (99m)Tc MDP scintigraphy in all 22 patients with skeletal lesions on both scans and over (18)F FDG PET/CT in 11 of 16 patients with skeletal metastases on (18)F FDG PET/CT. In two patients, (18)F NaF PET/CT showed skeletal metastases not seen on either of the other two scans. Extraskeletal lesions were identified by (18)F FDG PET/CT in 28 of 52 subjects.Our prospective pilot-phase trial demonstrates superior image quality and evaluation of skeletal disease extent with (18)F NaF PET/CT over (99m)Tc MDP scintigraphy and (18)F FDG PET/CT. At the same time, (18)F FDG PET detects extraskeletal disease that can significantly change disease management. As such, a combination of (18)F FDG PET/CT and (18)F NaF PET/CT may be necessary for cancer detection. Additional evaluation with larger cohorts is required to confirm these preliminary findings.

  View details for DOI 10.1007/s11307-011-0486-2

  View details for Web of Science ID 000301584100013

  View details for PubMedID 21479710

• Fiber-based system for imaging tumor margins with Cerenkov Luminescence Liu, H., Carpenter, C. M., Jiang, H., Pratx, G., Sun, C., Buchin, M. P., Gambhir, S. S., Xing, L., Cheng, Z. AMER CHEMICAL SOC. 2012

  View details for Web of Science ID 000324503202159

• Immunomodulation of Curcumin on Adoptive Therapy with T Cell Functional Imaging in Mice CANCER PREVENTION RESEARCH Chang, Y., Chuang, H., Hsu, C., Liu, R., Gambhir, S. S., Hwang, J. 2012; 5 (3): 444-452

  Abstract

  Adoptive T-cell therapy involves the ex vivo expansion and subsequent transfusion of tumor-specific T lymphocytes to eliminate tumors. Using immune modulators to block immunosuppressive factors in the tumor microenvironment has emerged as a promising strategy to enhance T-cell-mediated tumor regression. Curcumin, a major component of turmeric, has been shown to possess antitumor and immunomodulatory effects by regulating a diverse range of molecular targets. Thus, we hypothesize that these beneficial effects of curcumin may improve the therapeutic efficacy of adoptive therapy. Here, we have shown that curcumin enhances cytotoxicity of CD8(+) T cells toward tumors via alteration of the tumor microenvironment when combined with adoptive therapy. We found that T-cell accumulation and function were increased in combined treatment due to the blockade of different immunosuppressors, including TGF-β, indoleamine 2,3-dioxygenase, and regulatory T cells. Furthermore, bioluminescent imaging with a granzyme B promoter-conjugated optical reporter also reflected improved cytotoxicity of antigen-specific CD8(+) T cells in tumor-bearing mice during treatment. These findings suggest that combination of multitargeting drugs, such as curcumin, with adoptive therapy may have potential for clinical application. In addition, using a granzyme B-specific imaging reporter to assess T-cell function may also be applied for the development and therapeutic evaluation of new immunotherapy in preclinical studies.

  View details for DOI 10.1158/1940-6207.CAPR-11-0308

  View details for Web of Science ID 000300987800011

  View details for PubMedID 22135043

• Placental sFlt-1 Production Is Essential for Normal Pregnancy: Relevance to the Mechanisms of Preeclampsia. Nayak, N. R., Ambati, B. K., Dhal, S., Druzin, M. L., Gambhir, S. S., Fan, X. SAGE PUBLICATIONS INC. 2012: 86A

  View details for Web of Science ID 000329543601046

• Optical Imaging with Her2-Targeted Affibody Molecules Can Monitor Hsp90 Treatment Response in a Breast Cancer Xenograft Mouse Model CLINICAL CANCER RESEARCH van de Ven, S. M., Elias, S. G., Chan, C. T., Miao, Z., Cheng, Z., De, A., Gambhir, S. S. 2012; 18 (4): 1073-1081

  Abstract

  To determine whether optical imaging can be used for in vivo therapy response monitoring as an alternative to radionuclide techniques. For this, we evaluated the known Her2 response to 17-dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride (17-DMAG) treatment, an Hsp90 inhibitor.After in vitro 17-DMAG treatment response evaluation of MCF7 parental cells and 2 HER2-transfected clones (clone A medium, B high Her2 expression), we established human breast cancer xenografts in nude mice (only parental and clone B) for in vivo evaluation. Mice received 120 mg/kg of 17-DMAG in 4 doses at 12-hour intervals intraperitonially (n = 14) or PBS as carrier control (n = 9). Optical images were obtained both pretreatment (day 0) and posttreatment (day 3, 6, and 9), always 5 hours postinjection of 500 pmol of anti-Her2 Affibody-AlexaFluor680 via tail vein (with preinjection background subtraction). Days 3 and 9 in vivo optical imaging signal was further correlated with ex vivo Her2 levels by Western blot after sacrifice.Her2 expression decreased with 17-DMAG dose in vitro. In vivo optical imaging signal was reduced by 22.5% in clone B (P = 0.003) and by 9% in MCF7 parental tumors (P = 0.23) 3 days after 17-DMAG treatment; optical imaging signal recovered in both tumor types at days 6 to 9. In the carrier group, no signal reduction was observed. Pearson correlation of in vivo optical imaging signal with ex vivo Her2 levels ranged from 0.73 to 0.89.Optical imaging with an affibody can be used to noninvasively monitor changes in Her2 expression in vivo as a response to treatment with an Hsp90 inhibitor, with results similar to response measurements in positron emission tomography imaging studies.

  View details for DOI 10.1158/1078-0432.CCR-10-3213

  View details for Web of Science ID 000300628100017

  View details for PubMedID 22235098

  View details for PubMedCentralID PMC3288571

• First Experience with Clinical-Grade [F-18]FPP(RGD)(2): An Automated Multi-step Radiosynthesis for Clinical PET Studies MOLECULAR IMAGING AND BIOLOGY Chin, F. T., Shen, B., Liu, S., Berganos, R. A., Chang, E., Mittra, E., Chen, X., Gambhir, S. S. 2012; 14 (1): 88-95

  Abstract

  A reliable and routine process to introduce a new ¹⁸F-labeled dimeric RGD-peptide tracer ([¹⁸F]FPP(RGD₂) for noninvasive imaging of α(v)β₃ expression in tumors needed to be developed so the tracer could be evaluated for the first time in man. Clinical-grade [¹⁸F]FPP(RGD)₂ was screened in mouse prior to our first pilot study in human.[¹⁸F]FPP(RGD)₂ was synthesized by coupling 4-nitrophenyl-2-[¹⁸F]fluoropropionate ([¹⁸F]NPE) with the dimeric RGD-peptide (PEG₃-c(RGDyK)₂). Imaging studies with [¹⁸F]FPP(RGD)₂ in normal mice and a healthy human volunteer were carried out using small animal and clinical PET scanners, respectively.Through optimization of each radiosynthetic step, [¹⁸F]FPP(RGD)₂ was obtained with RCYs of 16.9 ± 2.7% (n = 8, EOB) and specific radioactivity of 114 ± 72 GBq/μmol (3.08 ± 1.95 Ci/μmol; n = 8, EOB) after 170 min of radiosynthesis. In our mouse studies, high radioactivity uptake was only observed in the kidneys and bladder with the clinical-grade tracer. Favorable [¹⁸F]FPP(RGD)₂ biodistribution in human studies, with low background signal in the head, neck, and thorax, showed the potential applications of this RGD-peptide tracer for detecting and monitoring tumor growth and metastasis.A reliable, routine, and automated radiosynthesis of clinical-grade [¹⁸F]FPP(RGD)₂ was established. PET imaging in a healthy human volunteer illustrates that [¹⁸F]FPP(RGD)₂ possesses desirable pharmacokinetic properties for clinical noninvasive imaging of α(v)β₃ expression. Further imaging studies using [¹⁸F]FPP(RGD)₂ in patient volunteers are now under active investigation.

  View details for DOI 10.1007/s11307-011-0477-3

  View details for Web of Science ID 000301583900012

  View details for PubMedID 21400112

  View details for PubMedCentralID PMC3617483

• Prospective comparison of combined F-18-FDG and F-18-NaF PET/CT vs. F-18-FDG PET/CT imaging for detection of malignancy EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Lin, F. I., Rao, J. E., Mittra, E. S., Nallapareddy, K., Chengapa, A., Dick, D. W., Gambhir, S. S., Iagaru, A. 2012; 39 (2): 262-270

  Abstract

  Typically, (18)F-FDG PET/CT and (18)F-NaF PET/CT scans are done as two separate studies on different days to allow sufficient time for the radiopharmaceutical from the first study to decay. This is inconvenient for the patients and exposes them to two doses of radiation from the CT component of the examinations. In the current study, we compared the clinical usefulness of a combined (18)F-FDG/(18)F-NaF PET/CT scan with that of a separate (18)F-FDG-only PET/CT scan.There were 62 patients enrolled in this prospective trial. All had both an (18)F-FDG-alone PET/CT scan and a combined (18)F-FDG/(18)F-NaF PET/CT scan. Of the 62 patients, 53 (85%) received simultaneous tracer injections, while 9 (15%) received (18)F-NaF subsequent to the initial (18)F-FDG dose (average delay 2.2 h). Images were independently reviewed for PET findings by two Board-Certified nuclear medicine physicians, with discrepancies resolved by a third reader. Interpreters were instructed to only report findings that were concerning for malignancy. Reading the (18)F-FDG-only scan first for half of the patients controlled for order bias.In 15 of the 62 patients (24%) neither the (18)F-FDG-only PET/CT scan nor the combined (18)F-FDG/(18)F-NaF PET/CT scan identified malignancy. In the remaining 47 patients who had PET findings of malignancy, a greater number of lesions were detected in 16 of 47 patients (34%) using the combined (18)F-FDG/(18)F-NaF PET/CT scan compared to the (18)F-FDG-only PET/CT scan. In 2 of these 47 patients (4%), the (18)F-FDG-only scan demonstrated soft tissue lesions that were not prospectively identified on the combined study. In 29 of these 47 patients (62%), the combined scan detected an equal number of lesions compared to the (18)F-FDG-only scan. Overall, 60 of all the 62 patients (97%) showed an equal or greater number of lesions on the combined scan than on the (18)F-FDG-only scan.The current study demonstrated that (18)F-FDG and (18)F-NaF can be combined in a single PET/CT scan by administering the two radiopharmaceuticals simultaneously or in sequence on the same day. In addition to patient convenience and reduced radiation exposure from the CT component, the combined (18)F-FDG/(18)F-NaF PET/CT scan appeared to increase the sensitivity for detection of osseous lesions compared to the (18)F-FDG-only PET/CT scan in the studied population.

  View details for DOI 10.1007/s00259-011-1971-1

  View details for Web of Science ID 000302286600009

  View details for PubMedID 22065013

• Proof-of-Concept Study of Monitoring Cancer Drug Therapy with Cerenkov Luminescence Imaging JOURNAL OF NUCLEAR MEDICINE Xu, Y., Chang, E., Liu, H., Jiang, H., Gambhir, S. S., Cheng, Z. 2012; 53 (2): 312-317

  Abstract

  Cerenkov luminescence imaging (CLI) has emerged as a less expensive, easier-to-use, and higher-throughput alternative to other nuclear imaging modalities such as PET. It is expected that CLI will find many applications in biomedical research such as cancer detection, probe development, drug screening, and therapy monitoring. In this study, we explored the possibility of using CLI to monitor drug efficacy by comparisons against PET. To assess the performance of both modalities in therapy monitoring, 2 murine tumor models (large cell lung cancer cell line H460 and prostate cancer cell line PC3) were given bevacizumab versus vehicle treatments. Two common radiotracers, 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) and (18)F-FDG, were used to monitor bevacizumab treatment efficacy.One group of mice (n = 6) was implanted with H460 xenografts bilaterally in the shoulder region, divided into treatment and control groups (n = 3 each), injected with (18)F-FLT, and imaged with PET immediately followed by CLI. The other group of mice (n = 6) was implanted with PC3 xenografts in the same locations, divided into treatment and control groups (n = 3 each), injected with (18)F-FDG, and imaged by the same modalities. Bevacizumab treatment was performed by 2 injections of 20 mg/kg at days 0 and 2.On (18)F-FLT scans, both CLI and PET revealed significantly decreased signals from H460 xenografts in treated mice from pretreatment to day 3. Moderately increased to unchanged signals were observed in untreated mice. On (18)F-FDG scans, both CLI and PET showed relatively unchanged signals from PC3 tumors in both treated and control groups. Quantifications of tumor signals of Cerenkov luminescence and PET images showed that the 2 modalities had excellent correlations (R(2) > 0.88 across all study groups).CLI and PET exhibit excellent correlations across different tumor xenografts and radiotracers. This is the first study, to our knowledge, demonstrating the use of CLI for monitoring cancer treatment. The findings warrant further exploration and optimization of CLI as an alternative to PET in preclinical therapeutic monitoring and drug screening.

  View details for DOI 10.2967/jnumed.111.094623

  View details for Web of Science ID 000300032800024

  View details for PubMedID 22241909

• Pharmacokinetically Stabilized Cystine Knot Peptides That Bind Alpha-v-Beta-6 Integrin with Single-Digit Nanomolar Affinities for Detection of Pancreatic Cancer CLINICAL CANCER RESEARCH Kimura, R. H., Teed, R., Hackel, B. J., Pysz, M. A., Chuang, C. Z., Sathirachinda, A., Willmann, J. K., Gambhir, S. S. 2012; 18 (3): 839-849

  Abstract

  Detection of pancreatic cancer remains a high priority and effective diagnostic tools are needed for clinical applications. Many cancer cells overexpress integrin α(v)β(6), a cell surface receptor being evaluated as a novel clinical biomarker.To validate this molecular target, several highly stable cystine knot peptides were engineered by directed evolution to bind specifically and with high affinity (3-6 nmol/L) to integrin α(v)β(6). The binders do not cross-react with related integrin α(v)β(5), integrin α(5)β(1), or tumor-angiogenesis-associated integrin, α(v)β(3).Positron emission tomography showed that these disulfide-stabilized peptides rapidly accumulate at tumors expressing integrin α(v)β(6). Clinically relevant tumor-to-muscle ratios of 7.7 ± 2.4 to 11.3 ± 3.0 were achieved within 1 hour after radiotracer injection. Minimization of off-target dosing was achieved by reformatting α(v)β(6)-binding activities across various natural and pharmacokinetically stabilized cystine knot scaffolds with different amino acid content. We show that the primary sequence of a peptide scaffold directs its pharmacokinetics. Scaffolds with high arginine or glutamic acid content suffered high renal retention of more than 75% injected dose per gram (%ID/g). Substitution of these amino acids with renally cleared amino acids, notably serine, led to significant decreases in renal accumulation of less than 20%ID/g 1 hour postinjection (P < 0.05, n = 3).We have engineered highly stable cystine knot peptides with potent and specific integrin α(v)β(6)-binding activities for cancer detection. Pharmacokinetic engineering of scaffold primary sequence led to significant decreases in off-target radiotracer accumulation. Optimization of binding affinity, specificity, stability, and pharmacokinetics will facilitate translation of cystine knots for cancer molecular imaging.

  View details for DOI 10.1158/1078-0432.CCR-11-1116

  View details for Web of Science ID 000300115000027

  View details for PubMedID 22173551

  View details for PubMedCentralID PMC3271184

• A Novel Clinically Translatable Fluorescent Nanoparticle for Targeted Molecular Imaging of Tumors in Living Subjects NANO LETTERS Gao, J., Chen, K., Luong, R., Bouley, D. M., Mao, H., Qiao, T., Gambhir, S. S., Cheng, Z. 2012; 12 (1): 281-286

  Abstract

  The use of quantum dots (QDs) in biomedical research has grown tremendously, yet successful examples of clinical applications are absent due to many clinical concerns. Here, we report on a new type of stable and biocompatible dendron-coated InP/ZnS core/shell QD as a clinically translatable nanoprobe for molecular imaging applications. The QDs (QD710-Dendron) were demonstrated to hold several significant features: near-infrared (NIR) emission, high stability in biological media, suitable size with possible renal clearance, and ability of extravasation. More importantly, a pilot mouse toxicity study confirmed that QD710-Dendron lacks significant toxicity at the doses tested. The acute tumor uptake of QD710-Dendron resulted in good contrast from the surrounding nontumorous tissues, indicating the possibility of passive targeting of the QDs. The highly specific targeting of QD710-Dendron-RGD(2) to integrin α(v)β(3)-positive tumor cells resulted in high tumor uptake and long retention of the nanoprobe at tumor sites. In summary, QD710-Dendron and RGD-modified nanoparticles demonstrate small size, high stability, biocompatibility, favorable in vivo pharmacokinetics, and successful tumor imaging properties. These features satisfy the requirements for clinical translation and should promote efforts to further investigate the possibility of using QD710-Dendron-based nanoprobes in the clinical setting in the near future.

  View details for DOI 10.1021/nl203526f

  View details for Web of Science ID 000298943100049

  View details for PubMedID 22172022

  View details for PubMedCentralID PMC3256290

• Imaging Techniques in Drug Development and Clinical Practice DRUG DELIVERY IN ONCOLOGY: FROM BASIC RESEARCH TO CANCER THERAPY, VOLS 1-3 Chang, J. C., Gambhir, S. S., Willmann, J. K., Kratz, F., Senter, P., Steinhagen, H. 2012: 189–224

  View details for Web of Science ID 000338009100010

• Photoacoustic Imaging Using a 9F MicroLinear CMUT ICE Catheter IEEE International Ultrasonics Symposium (IUS) Nikoozadeh, A., Choe, J. W., Kothapalli, S., Moini, A., Sanjani, S. S., Kamaya, A., Oralkan, O., Gambhir, S. S., Khuri-Yakub, P. T. IEEE. 2012: 24–27

  View details for DOI 10.1109/ULTSYM.2012.0007

  View details for Web of Science ID 000326960200006

• Response to Intra-Arterial Oncolytic Virotherapy with the Herpes Virus NV1020 Evaluated by [F-18]Fluorodeoxyglucose Positron Emission Tomography and Computed Tomography HUMAN GENE THERAPY Sze, D. Y., Iagaru, A. H., Gambhir, S. S., de Haan, H. A., Reid, T. R. 2012; 23 (1): 91-97

  Abstract

  Oncolytic virotherapy poses unique challenges to the evaluation of tumor response. We hypothesized that the addition of [(18)F]fluorodeoxyglucose (FDG) positron emission tomography (PET) to standard computed tomography (CT) evaluation would improve diagnostic and prognostic power of the measurement of tumor response to oncolytic virotherapy. A phase I/II trial was conducted to investigate treatment of hepatic metastases from colorectal carcinoma using intra-arterial administration of the oncolytic herpes virus NV1020. Both contrast-enhanced CT and FDG PET were obtained on each patient at each time point. Quantitative FDG PET and CT responses were correlated with each other and with clinical outcome metrics. A majority of patients showed initial post-viral infusion increases in tumor size (69%) or in standardized uptake value (SUV) (80%) large enough to qualify as progressive disease. Most showed subsequent decreases in tumor size (64%) or SUV (83%) enough to be reclassified as partial response or stable disease. Late PET and CT imaging results correlated well with each other and with clinical outcomes, but results from early in the treatment scheme did not correlate with each other, with later results, or with clinical outcomes. The addition of FDG PET to the evaluation of tumor response to the oncolytic virus NV1020 did not provide useful diagnostic or prognostic data. More sophisticated molecular imaging will need to be developed to monitor the effects of this novel class of antineoplastic agents.

  View details for DOI 10.1089/hum.2011.141

  View details for Web of Science ID 000299604000011

  View details for PubMedID 21895536

• Raman's "Effect" on Molecular Imaging JOURNAL OF NUCLEAR MEDICINE Zavaleta, C. L., Kircher, M. F., Gambhir, S. S. 2011; 52 (12): 1839-1844

  Abstract

  Raman spectroscopy is an optical technique that offers unsurpassed sensitivity and multiplexing capabilities to the field of molecular imaging. In the past, Raman spectroscopy had predominantly been used as an analytic tool for routine chemical analysis, but more recently, researchers have been able to harness its unique properties for imaging and spectral analysis of molecular interactions in cell populations and preclinical animal models. Additionally, researchers have already begun to translate this optical technique into a novel clinical diagnostic tool using various endoscopic strategies.

  View details for DOI 10.2967/jnumed.111.087775

  View details for Web of Science ID 000298162500016

  View details for PubMedID 21868625

• In Vitro and in Vivo Molecular Imaging of Estrogen Receptor alpha and beta Homo- and Heterodimerization: Exploration of New Modes of Receptor Regulation MOLECULAR ENDOCRINOLOGY Paulmurugan, R., Tamrazi, A., Massoud, T. F., Katzenellenbogen, J. A., Gambhir, S. S. 2011; 25 (12): 2029-2040

  Abstract

  Estrogen receptor (ER) biology reflects the actions of estrogens through the two receptors, ERα and ERβ, although little is known regarding the preference for formation of ER homo- vs. heterodimers, and how this is affected by the level of ligand occupancy and preferential ligand affinity for one of the ER subtypes. In this report, we use a split optical reporter-protein complementation system to demonstrate the physical interaction between ERα and ERβ in response to different ER ligands in cells and, for the first time, by in vivo imaging in living animals. The genetically encoded reporter vectors constructed with the ligand-binding domains of ERα and ERβ, fused to split firefly or Renilla luciferase (Fluc or hRluc) fragments, were used for this study. This molecular proteomic technique was used to detect ERα/ERα or ERβ/ERβ homodimerization, or ERα/ERβ heterodimerization induced by ER subtype-selective and nonselective ligands, and selective ER modulators (SERM), as well as in dimers in which one mutant monomer was unable to bind estradiol. The SERM-bound ERα and ERβ form the strongest dimers, and subtype-preferential homodimerization was seen with ERα-selective ligands (methyl piperidino pyrazole/propyl pyrazole triol) and the ERβ-selective ligands (diarylpropionitrile/tetrahydrochrysene/genistein). We also demonstrated that a single ligand-bound monomer can form homo- or heterodimers with an apo-monomer. Xenografts of human embryonic kidney 293T cells imaged in living mice by bioluminescence showed real-time ligand induction of ERα/ERβ heterodimerization and reversal of dimerization upon ligand withdrawal. The results from this study demonstrate the value of the split luciferase-based complementation system for studying ER-subtype interactions in cells and for evaluating them in living animals by noninvasive imaging. They also probe what combinations of ERα and ERβ dimers might be the mediators of the effects of different types of ER ligands given at different doses.

  View details for DOI 10.1210/me.2011-1145

  View details for PubMedID 22052998

• Non-invasive Bioluminescence Imaging of Myoblast-Mediated Hypoxia-Inducible Factor-1 Alpha Gene Transfer MOLECULAR IMAGING AND BIOLOGY Gheysens, O., Chen, I. Y., Rodriguez-Porcel, M., Chan, C., Rasooly, J., Vaerenberg, C., Paulmurugan, R., Willmann, J. K., Deroose, C., Wu, J., Gambhir, S. S. 2011; 13 (6): 1124-1132

  Abstract

  We tested a novel imaging strategy, in which both the survival of transplanted myoblasts and their therapeutic transgene expression, a recombinant hypoxia-inducible factor-1α (HIF-1α-VP2), can be monitored using firefly luciferase (fluc) and Renilla luciferase (hrl) bioluminescence reporter genes, respectively.The plasmid pUbi-hrl-pUbi-HIF-1α-VP2, which expresses both hrl and HIF-1α-VP2 using two ubiquitin promoters, was characterized in vitro. C2c12 myoblasts stably expressing fluc and transiently transfected with pUbi-hrl-pUbi-HIF-1α-VP2 were injected into the mouse hindlimb. Both hrl and fluc expression were monitored using bioluminescence imaging (BLI).Strong correlations existed between the expression of hRL and each of HIF-1α-VP2, VEGF, and PlGF (r(2) > 0.83, r(2) > 0.82, and r(2) > 0.97, respectively). In vivo, both transplanted cells and HIF-1α-VP2 transgene expression were successfully imaged using BLI.An objective evaluation of myoblast-mediated gene transfer in living mice can be performed by monitoring both the survival and the transgene expression of transplanted myoblasts using the techniques developed herein.

  View details for DOI 10.1007/s11307-011-0471-9

  View details for Web of Science ID 000296794400009

  View details for PubMedID 21267661

  View details for PubMedCentralID PMC4657136

• Non-Invasive Imaging of Cysteine Cathepsin Activity in Solid Tumors Using a Cu-64-Labeled Activity-Based Probe PLOS ONE Ren, G., Blum, G., Verdoes, M., Liu, H., Syed, S., Edgington, L. E., Gheysens, O., Miao, Z., Jiang, H., Gambhir, S. S., Bogyo, M., Cheng, Z. 2011; 6 (11)

  Abstract

  The papain family of cysteine cathepsins are actively involved in multiple stages of tumorigenesis. Because elevated cathepsin activity can be found in many types of human cancers, they are promising biomarkers that can be used to target radiological contrast agents for tumor detection. However, currently there are no radiological imaging agents available for these important molecular targets. We report here the development of positron emission tomography (PET) radionuclide-labeled probes that target the cysteine cathepsins by formation of an enzyme activity-dependent bond with the active site cysteine. These probes contain an acyloxymethyl ketone (AOMK) functional group that irreversibly labels the active site cysteine of papain family proteases attached to a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) tag for labeling with (64)Cu for PET imaging studies. We performed biodistribution and microPET imaging studies in nude mice bearing subcutaneous tumors expressing various levels of cysteine cathepsin activity and found that the extent of probe uptake by tumors correlated with overall protease activity as measured by biochemical methods. Furthermore, probe signals could be reduced by pre-treatment with a general cathepsin inhibitor. We also found that inclusion of a Cy5 tag on the probe increased tumor uptake relative to probes lacking this fluorogenic dye. Overall, these results demonstrate that small molecule activity-based probes carrying radio-tracers can be used to image protease activity in living subjects.

  View details for DOI 10.1371/journal.pone.0028029

  View details for Web of Science ID 000297789900039

  View details for PubMedID 22132198

  View details for PubMedCentralID PMC3221694

• Mathematical Model Identifies Blood Biomarker-Based Early Cancer Detection Strategies and Limitations SCIENCE TRANSLATIONAL MEDICINE Hori, S. S., Gambhir, S. S. 2011; 3 (109)

  Abstract

  Most clinical blood biomarkers lack the necessary sensitivity and specificity to reliably detect cancer at an early stage, when it is best treatable. It is not yet clear how early a clinical blood assay can be used to detect cancer or how biomarker-based strategies can be improved to enable earlier detection of smaller tumors. To address these issues, we developed a mathematical model describing dynamic plasma biomarker kinetics in relation to the growth of a tumor, beginning with a single cancer cell. To exemplify a realistic scenario in which biomarker is shed by both cancerous and noncancerous cells, we primed the model on ovarian tumor growth and CA125 shedding data, for which tumor growth parameters and shedding rates are readily available in published literature. We found that a tumor could grow unnoticed for more than 10.1 years and reach a volume of about π/6(25.36 mm)(3), corresponding to a spherical diameter of about 25.36 mm, before becoming detectable by current clinical blood assays. Model parameters were perturbed over log orders of magnitude to quantify ideal shedding rates and identify other blood-based strategies required for early submillimeter tumor detectability. The detection times we estimated are consistent with recently published tumor progression time lines based on clinical genomic sequencing data for several cancers. Here, we rigorously showed that shedding rates of current clinical blood biomarkers are likely 10(4)-fold too low to enable detection of a developing tumor within the first decade of tumor growth. The model presented here can be extended to virtually any solid cancer and associated biomarkers.

  View details for DOI 10.1126/scitranslmed.3003110

  View details for Web of Science ID 000297218300004

  View details for PubMedID 22089452

  View details for PubMedCentralID PMC3423335

• GLUT 5 Is Not Over-Expressed in Breast Cancer Cells and Patient Breast Cancer Tissues PLOS ONE Gowrishankar, G., Zitzmann-Kolbe, S., Junutula, A., Reeves, R., Levi, J., Srinivasan, A., Bruus-Jensen, K., Cyr, J., Dinkelborg, L., Gambhir, S. S. 2011; 6 (11)

  Abstract

  F18 2-Fluoro 2-deoxyglucose (FDG) has been the gold standard in positron emission tomography (PET) oncologic imaging since its introduction into the clinics several years ago. Seeking to complement FDG in the diagnosis of breast cancer using radio labeled fructose based analogs, we investigated the expression of the chief fructose transporter-GLUT 5 in breast cancer cells and human tissues. Our results indicate that GLUT 5 is not over-expressed in breast cancer tissues as assessed by an extensive immunohistochemistry study. RT-PCR studies showed that the GLUT 5 mRNA was present at minimal amounts in breast cancer cell lines. Further knocking down the expression of GLUT 5 in breast cancer cells using RNA interference did not affect the fructose uptake in these cell lines. Taken together these results are consistent with GLUT 5 not being essential for fructose uptake in breast cancer cells and tissues.

  View details for DOI 10.1371/journal.pone.0026902

  View details for Web of Science ID 000297154900052

  View details for PubMedID 22073218

  View details for PubMedCentralID PMC3206880

• A novel F-18-labeled two-helix scaffold protein for PET imaging of HER2-positive tumor EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Miao, Z., Ren, G., Jiang, L., Liu, H., Webster, J. M., Zhang, R., Namavari, M., Gambhir, S. S., Syud, F., Cheng, Z. 2011; 38 (11): 1977-1984

  Abstract

  Two-helix scaffold proteins (~ 5 kDa) against human epidermal growth factor receptor type 2 (HER2) have been discovered in our previous work. In this research we aimed to develop an (18)F-labeled two-helix scaffold protein for positron emission tomography (PET) imaging of HER2-positive tumors.An aminooxy-functionalized two-helix peptide (AO-MUT-DS) with high HER2 binding affinity was synthesized through conventional solid phase peptide synthesis. The purified linear peptide was cyclized by I(2) oxidation to form a disulfide bridge. The cyclic peptide was then conjugated with a radiofluorination synthon, 4-(18)F-fluorobenzyl aldehyde ((18)F-FBA), through the aminooxy functional group at the peptide N terminus (30% yield, non-decay corrected). The binding affinities of the peptides were analyzed by Biacore analysis. Cell uptake assay of the resulting PET probe, (18)F-FBO-MUT-DS, was performed at 37°C. (18)F-FBO-MUT-DS with high specific activity (20-32 MBq/nmol, 88-140 μCi/μg, end of synthesis) was injected into mice xenograft model bearing SKOV3 tumor. MicroPET and biodistribution and metabolic stability studies were then conducted.Cell uptake assays showed high and specific cell uptake (~12% applied activity at 1 h) by incubation of (18)F-FBO-MUT-DS with HER2 high-expressing SKOV3 ovarian cancer cells. The affinities (K(D)) of AO-MUT-DS and FBO-MUT-DS as tested by Biacore analysis were 2 and 1 nM, respectively. In vivo small animal PET demonstrated fast tumor targeting, high tumor accumulation, and good tumor to normal tissue contrast of (18)F-FBO-MUT-DS. Biodistribution studies further revealed that the probe had excellent tumor uptake (6.9%ID/g at 1 h post-injection) and was cleared through both liver and kidneys. Co-injection of the probe with 500 μg of HER2 Affibody protein reduced the tumor uptake (6.9 vs 1.8%ID/g, p < 0.05).F-FBO-MUT-DS displays excellent HER2 targeting ability and tumor PET imaging quality. The two-helix scaffold proteins are suitable for development of (18)F-based PET probes.

  View details for DOI 10.1007/s00259-011-1879-9

  View details for Web of Science ID 000295680200004

  View details for PubMedID 21761266

• Noninvasive cell-tracking methods NATURE REVIEWS CLINICAL ONCOLOGY Kircher, M. F., Gambhir, S. S., Grimm, J. 2011; 8 (11): 677-688

  Abstract

  Cell-based therapies, such as adoptive immunotherapy and stem-cell therapy, have received considerable attention as novel therapeutics in oncological research and clinical practice. The development of effective therapeutic strategies using tumor-targeted cells requires the ability to determine in vivo the location, distribution, and long-term viability of the therapeutic cell populations as well as their biological fate with respect to cell activation and differentiation. In conjunction with various noninvasive imaging modalities, cell-labeling methods, such as exogenous labeling or transfection with a reporter gene, allow visualization of labeled cells in vivo in real time, as well as monitoring and quantifying cell accumulation and function. Such cell-tracking methods also have an important role in basic cancer research, where they serve to elucidate novel biological mechanisms. In this Review, we describe the basic principles of cell-tracking methods, explain various approaches to cell tracking, and highlight recent examples for the application of such methods in animals and humans.

  View details for DOI 10.1038/nrclinonc.2011.141

  View details for Web of Science ID 000296812500009

  View details for PubMedID 21946842

• Endothelial Cells Derived From Human iPSCS Increase Capillary Density and Improve Perfusion in a Mouse Model of Peripheral Arterial Disease ARTERIOSCLEROSIS THROMBOSIS AND VASCULAR BIOLOGY Rufaihah, A. J., Huang, N. F., Jame, S., Lee, J. C., Nguyen, H. N., Byers, B., De, A., Okogbaa, J., Rollins, M., Reijo-Pera, R., Gambhir, S. S., Cooke, J. P. 2011; 31 (11): E72-U44

  Abstract

  Stem cell therapy for angiogenesis and vascular regeneration has been investigated using adult or embryonic stem cells. In the present study, we investigated the potential of endothelial cells (ECs) derived from human induced pluripotent stem cells (hiPSCs) to promote the perfusion of ischemic tissue in a murine model of peripheral arterial disease.Endothelial differentiation was initiated by culturing hiPSCs for 14 days in differentiation media supplemented with BMP-4 and vascular endothelial growth factor. The hiPSC-ECs exhibited endothelial characteristics by forming capillary-like structures in matrigel and incorporating acetylated-LDL. They stained positively for EC markers such as KDR, CD31, CD144, and eNOS. In vitro exposure of hiPSC-ECs to hypoxia resulted in increased expression of various angiogenic related cytokines and growth factors. hiPSC-ECs were stably transduced with a double fusion construct encoded by the ubiquitin promoter, firefly luciferase for bioluminescence imaging and green fluorescence protein for fluorescent detection. The hiPSC-ECs (5×10(5)) were delivered by intramuscular injection into the ischemic hindlimb of SCID mice at day 0 and again on day 7 after femoral artery ligation (n=8). Bioluminescence imaging showed that hiPSC-ECs survived in the ischemic limb for at least 2 weeks. In addition, laser Doppler imaging showed that the ratio of blood perfusion was increased by hiPSC-EC treatment by comparison to the saline-treated group (0.58±0.12 versus 0.44±0.04; P=0.005). The total number of capillaries in the ischemic limb of mice receiving hiPSC-EC injections was greater than those in the saline-treated group (1284±155 versus 797±206 capillaries/mm(2)) (P<0.002).This study is a first step toward development of a regenerative strategy for peripheral arterial disease based on the use of ECs derived from hiPSCs.

  View details for DOI 10.1161/ATVBAHA.111.230938

  View details for PubMedID 21836062

• Theranostic nanomedicine. Accounts of chemical research Chen, X., Gambhir, S. S., Cheon, J. 2011; 44 (10): 841-?

  View details for DOI 10.1021/ar200231d

  View details for PubMedID 22004477

• Immobilizing Reporters for Molecular Imaging of the Extracellular Microenvironment in Living Animals ACS CHEMICAL BIOLOGY Xia, Z., Xing, Y., Jeon, J., Kim, Y., Gall, J., Dragulescu-Andrasi, A., Gambhir, S. S., Rao, J. 2011; 6 (10): 1117-1126

  Abstract

  We report here an immobilization strategy using a collagen binding protein to deliver and confine synthetic reporters to the extracellular microenvironment in vivo for noninvasively imaging the activity of targets in the microenvironment. We show that the immobilization of reporters on collagens in the local microenvironment is highly efficient and physiologically stable for repetitive, long-term imaging. By using this strategy we successfully developed an immobilized bioluminescent activatable reporter and a dual-modality reporter to map and quantitatively image the activity of extracellular matrix metalloproteinases (MMP) in tumor-bearing mice. The inhibition of MMP activity by chemical inhibitor was also demonstrated in living subjects. We further demonstrated the general applicability of this immobilization strategy by imaging MMP activity at the inflammation site in a mouse model. Our results show that the in vivo immobilization of reporters can be used as a general strategy for probing the local extracellular microenvironment.

  View details for DOI 10.1021/cb200135e

  View details for Web of Science ID 000296208100018

  View details for PubMedID 21830814

  View details for PubMedCentralID PMC3199358

• Molecular Imaging with Theranostic Nanoparticles ACCOUNTS OF CHEMICAL RESEARCH Jokerst, J. V., Gambhir, S. S. 2011; 44 (10): 1050-1060

  Abstract

  Nanoparticles (NPs) offer diagnostic and therapeutic capabilities not available with small molecules or microscale tools. As the field of molecular imaging has emerged from the blending of molecular biology with medical imaging, NP imaging is increasingly common for both therapeutic and diagnostic applications. The term theranostic describes technology with concurrent and complementary diagnostic and therapeutic capabilities. Although NPs have been FDA-approved for clinical use as transport vehicles for nearly 15 years, full translation of their theranostic potential is incomplete. However, NPs have shown remarkable success in the areas of drug delivery and magnetic resonance imaging. Emerging applications include image-guided resection, optical/photoacoustic imaging in vivo, contrast-enhanced ultrasound, and thermoablative therapy. Diagnosis with NPs in molecular imaging involves the correlation of the signal with a phenotype. The location and intensity of NP signals emanating from a living subject indicate the disease area's size, stage, and biochemical signature. Therapy with NPs uses the image for resection or delivery of a small molecule or RNA therapeutic. Ablation of the affected area is also possible via heat or radioactivity. The ideal theranostic NP includes several features: (1) it selectively and rapidly accumulates in diseased tissue; (2) it reports biochemical and morphological characteristics of the area; (3) it delivers an effective therapeutic; and (4) it is safe and biodegrades with nontoxic byproducts. Such a system contains a central imaging core surrounded by small molecule therapeutics. The system targets via ligands such as IgG and is protected from immune scavengers by a cloak of protective polymer. Although no NP has achieved all of the above criteria, many NPs possess one or more of these features. While the most clinically translatable NPs have been used in the field of magnetic resonance imaging, other types in development are quickly becoming more biocompatible through methods that modify their toxicity and biodistribution profiles. In this Account, we describe diagnostic imaging and therapeutic uses of NPs. We propose and offer examples of five primary types of nanoparticles with concurrent diagnostic and therapeutic uses.

  View details for DOI 10.1021/ar200106e

  View details for Web of Science ID 000296682400022

  View details for PubMedID 21919457

  View details for PubMedCentralID PMC3196845

• Synthesis of 2 '-Deoxy-2 '-[F-18]Fluoro-9-beta-D-Arabinofuranosylguanine: a Novel Agent for Imaging T-Cell Activation with PET MOLECULAR IMAGING AND BIOLOGY Namavari, M., Chang, Y., Kusler, B., Yaghoubi, S., Mitchell, B. S., Gambhir, S. S. 2011; 13 (5): 812-818

  Abstract

  9-(β-D-Arabinofuranosyl)guanine (AraG) is a guanosine analog that has a proven efficacy in the treatment of T-cell lymphoblastic disease. To test the possibility of using a radiofluorinated AraG as an imaging agent, we have synthesized 2'-deoxy-2'-[(18)F]fluoro-9-β-D-arabinofuranosylguanine ([(18)F]F-AraG) and investigated its uptake in T cells.We have synthesized [(18)F]F-AraG via a direct fluorination of 2-N-acetyl-6-O-((4-nitrophenyl)ethyl)-9-(3',5'-di-O-trityl-2'-O-trifyl-β-D-ribofuranosyl)guanine with [(18)F]KF/K.2.2.2 in DMSO at 85°C for 45 min. [(18)F]F-AraG uptake in both a CCRF-CEM leukemia cell line (unactivated) and activated primary thymocytes was evaluated.We have successfully prepared [(18)F]F-AraG in 7-10% radiochemical yield (decay corrected) with a specific activity of 0.8-1.3 Ci/μmol. Preliminary cell uptake experiments showed that both a CCRF-CEM leukemia cell line and activated primary thymocytes take up the [(18)F]F-AraG.For the first time to the best of our knowledge, [(18)F]F-AraG has been successfully synthesized by direct fluorination of an appropriate precursor of a guanosine nucleoside. This approach maybe also useful for the synthesis of other important positron emission tomography (PET) probes such as [(18)F]FEAU, [(18)F]FMAU, and [(18)F]FBAU which are currently synthesized by multiple steps and involve lengthy purification. The cell uptake studies support future studies to investigate the use of [(18)F]F-AraG as a PET imaging agent of T cells.

  View details for DOI 10.1007/s11307-010-0414-x

  View details for PubMedID 20838911

• Gold Nanoparticles: A Revival in Precious Metal Administration to Patients NANO LETTERS Thakor, A. S., Jokerst, J., Zavaleta, C., Massoud, T. F., Gambhir, S. S. 2011; 11 (10): 4029-4036

  Abstract

  Gold has been used as a therapeutic agent to treat a wide variety of rheumatic diseases including psoriatic arthritis, juvenile arthritis, and discoid lupus erythematosus. Although the use of gold has been largely superseded by newer drugs, gold nanoparticles are being used effectively in laboratory based clinical diagnostic methods while concurrently showing great promise in vivo either as a diagnostic imaging agent or a therapeutic agent. For these reasons, gold nanoparticles are therefore well placed to enter mainstream clinical practice in the near future. Hence, the present review summarizes the chemistry, pharmacokinetics, biodistribution, metabolism, and toxicity of bulk gold in humans based on decades of clinical observation and experiments in which gold was used to treat patients with rheumatoid arthritis. The beneficial attributes of gold nanoparticles, such as their ease of synthesis, functionalization, and shape control are also highlighted demonstrating why gold nanoparticles are an attractive target for further development and optimization. The importance of controlling the size and shape of gold nanoparticles to minimize any potential toxic side effects is also discussed.

  View details for DOI 10.1021/nl202559p

  View details for PubMedID 21846107

• Preclinical Derivation and Imaging of Autologously Transplanted Canine Induced Pluripotent Stem Cells JOURNAL OF BIOLOGICAL CHEMISTRY Lee, A. S., Xu, D., Plews, J. R., Nguyen, P. K., Nag, D., Lyons, J. K., Han, L., Hu, S., Lan, F., Liu, J., Huang, M., Narsinh, K. H., Long, C. T., de Almeida, P. E., Levi, B., Kooreman, N., Bangs, C., Pacharinsak, C., Ikeno, F., Yeung, A. C., Gambhir, S. S., Robbins, R. C., Longaker, M. T., Wu, J. C. 2011; 286 (37): 32697-32704

  Abstract

  Derivation of patient-specific induced pluripotent stem cells (iPSCs) opens a new avenue for future applications of regenerative medicine. However, before iPSCs can be used in a clinical setting, it is critical to validate their in vivo fate following autologous transplantation. Thus far, preclinical studies have been limited to small animals and have yet to be conducted in large animals that are physiologically more similar to humans. In this study, we report the first autologous transplantation of iPSCs in a large animal model through the generation of canine iPSCs (ciPSCs) from the canine adipose stromal cells and canine fibroblasts of adult mongrel dogs. We confirmed pluripotency of ciPSCs using the following techniques: (i) immunostaining and quantitative PCR for the presence of pluripotent and germ layer-specific markers in differentiated ciPSCs; (ii) microarray analysis that demonstrates similar gene expression profiles between ciPSCs and canine embryonic stem cells; (iii) teratoma formation assays; and (iv) karyotyping for genomic stability. Fate of ciPSCs autologously transplanted to the canine heart was tracked in vivo using clinical positron emission tomography, computed tomography, and magnetic resonance imaging. To demonstrate clinical potential of ciPSCs to treat models of injury, we generated endothelial cells (ciPSC-ECs) and used these cells to treat immunodeficient murine models of myocardial infarction and hindlimb ischemia.

  View details for DOI 10.1074/jbc.M111.235739

  View details for Web of Science ID 000294726800078

  View details for PubMedID 21719696

  View details for PubMedCentralID PMC3173214

• Adipose tissue-derived stem cells display a proangiogenic phenotype on 3D scaffolds. Journal of biomedical materials research. Part A Neofytou, E. A., Chang, E., Patlola, B., Joubert, L., Rajadas, J., Gambhir, S. S., Cheng, Z., Robbins, R. C., Beygui, R. E. 2011; 98 (3): 383-393

  Abstract

  Ischemic heart disease is the leading cause of death worldwide. Recent studies suggest that adipose tissue-derived stem cells (ASCs) can be used as a potential source for cardiovascular tissue engineering due to their ability to differentiate along the cardiovascular lineage and to adopt a proangiogenic phenotype. To understand better ASCs' biology, we used a novel 3D culture device. ASCs' and b.END-3 endothelial cell proliferation, migration, and vessel morphogenesis were significantly enhanced compared to 2D culturing techniques. ASCs were isolated from inguinal fat pads of 6-week-old GFP+/BLI+ mice. Early passage ASCs cells (P3-P4), PKH26-labeled murine b.END-3 cells or a co-culture of ASCs and b.END-3 cells were seeded at a density of 1 × 10(5) on three different surface configurations: (a) a 2D surface of tissue culture plastic, (b) Matrigel, and (c) a highly porous 3D scaffold fabricated from inert polystyrene. VEGF expression, cell proliferation, and tubulization, were assessed using optical microscopy, fluorescence microscopy, 3D confocal microscopy, and SEM imaging (n = 6). Increased VEGF levels were seen in conditioned media harvested from co-cultures of ASCs and b.END-3 on either Matrigel or a 3D matrix. Fluorescence, confocal, SEM, bioluminescence revealed improved cell, proliferation, and tubule formation for cells seeded on the 3D polystyrene matrix. Collectively, these data demonstrate that co-culturing ASCs with endothelial cells in a 3D matrix environment enable us to generate prevascularized tissue-engineered constructs. This can potentially help us to surpass the tissue thickness limitations faced by the tissue engineering community today.

  View details for DOI 10.1002/jbm.a.33113

  View details for PubMedID 21630430

• Adipose tissue-derived stem cells display a proangiogenic phenotype on 3D scaffolds JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A Neofytou, E. A., Chang, E., Patloia, B., Joubert, L., Rajadas, J., Gambhir, S. S., Cheng, Z., Robbins, R. C., Beygui, R. E. 2011; 98A (3): 383-393

  Abstract

  Ischemic heart disease is the leading cause of death worldwide. Recent studies suggest that adipose tissue-derived stem cells (ASCs) can be used as a potential source for cardiovascular tissue engineering due to their ability to differentiate along the cardiovascular lineage and to adopt a proangiogenic phenotype. To understand better ASCs' biology, we used a novel 3D culture device. ASCs' and b.END-3 endothelial cell proliferation, migration, and vessel morphogenesis were significantly enhanced compared to 2D culturing techniques. ASCs were isolated from inguinal fat pads of 6-week-old GFP+/BLI+ mice. Early passage ASCs cells (P3-P4), PKH26-labeled murine b.END-3 cells or a co-culture of ASCs and b.END-3 cells were seeded at a density of 1 × 10(5) on three different surface configurations: (a) a 2D surface of tissue culture plastic, (b) Matrigel, and (c) a highly porous 3D scaffold fabricated from inert polystyrene. VEGF expression, cell proliferation, and tubulization, were assessed using optical microscopy, fluorescence microscopy, 3D confocal microscopy, and SEM imaging (n = 6). Increased VEGF levels were seen in conditioned media harvested from co-cultures of ASCs and b.END-3 on either Matrigel or a 3D matrix. Fluorescence, confocal, SEM, bioluminescence revealed improved cell, proliferation, and tubule formation for cells seeded on the 3D polystyrene matrix. Collectively, these data demonstrate that co-culturing ASCs with endothelial cells in a 3D matrix environment enable us to generate prevascularized tissue-engineered constructs. This can potentially help us to surpass the tissue thickness limitations faced by the tissue engineering community today.

  View details for DOI 10.1002/jbm.a.33113

  View details for Web of Science ID 000293699800007

• Advanced contrast nanoagents for photoacoustic molecular imaging, cytometry, blood test and photothermal theranostics CONTRAST MEDIA & MOLECULAR IMAGING de la Zerda, A., Kim, J., Galanzha, E. I., Gambhir, S. S., Zharov, V. P. 2011; 6 (5): 346-369

  Abstract

  Various nanoparticles have raised significant interest over the past decades for their unique physical and optical properties and biological utilities. Here we summarize the vast applications of advanced nanoparticles with a focus on carbon nanotube (CNT)-based or CNT-catalyzed contrast agents for photoacoustic (PA) imaging, cytometry and theranostics applications based on the photothermal (PT) effect. We briefly review the safety and potential toxicity of the PA/PT contrast nanoagents, while showing how the physical properties as well as multiple biological coatings change their toxicity profiles and contrasts. We provide general guidelines needed for the validation of a new molecular imaging agent in living subjects, and exemplify these guidelines with single-walled CNTs targeted to α(v) β(3) , an integrin associated with tumor angiogenesis, and golden carbon nanotubes targeted to LYVE-1, endothelial lymphatic receptors. An extensive review of the potential applications of advanced contrast agents is provided, including imaging of static targets such as tumor angiogenesis receptors, in vivo cytometry of dynamic targets such as circulating tumor cells and nanoparticles in blood, lymph, bones and plants, methods to enhance the PA and PT effects with transient and stationary bubble conjugates, PT/PA Raman imaging and multispectral histology. Finally, theranostic applications are reviewed, including the nanophotothermolysis of individual tumor cells and bacteria with clustered nanoparticles, nanothrombolysis of blood clots, detection and purging metastasis in sentinel lymph nodes, spectral hole burning and multiplex therapy with ultrasharp rainbow nanoparticles.

  View details for DOI 10.1002/cmmi.455

  View details for Web of Science ID 000300110400003

  View details for PubMedID 22025336

  View details for PubMedCentralID PMC4282188

• Preclinical Evaluation of Raman Nanoparticle Biodistribution for their Potential Use in Clinical Endoscopy Imaging SMALL Zavaleta, C. L., Hartman, K. B., Miao, Z., James, M. L., Kempen, P., Thakor, A. S., Nielsen, C. H., Sinclair, R., Cheng, Z., Gambhir, S. S. 2011; 7 (15): 2232-2240

  Abstract

  Raman imaging offers unsurpassed sensitivity and multiplexing capabilities. However, its limited depth of light penetration makes direct clinical translation challenging. Therefore, a more suitable way to harness its attributes in a clinical setting would be to couple Raman spectroscopy with endoscopy. The use of an accessory Raman endoscope in conjunction with topically administered tumor-targeting Raman nanoparticles during a routine colonoscopy could offer a new way to sensitively detect dysplastic lesions while circumventing Raman's limited depth of penetration and avoiding systemic toxicity. In this study, the natural biodistribution of gold surface-enhanced Raman scattering (SERS) nanoparticles is evaluated by radiolabeling them with (64) Cu and imaging their localization over time using micropositron emission tomography (PET). Mice are injected either intravenously (IV) or intrarectally (IR) with approximately 100 microcuries (μCi) (3.7 megabecquerel (MBq)) of (64) Cu-SERS nanoparticles and imaged with microPET at various time points post injection. Quantitative biodistribution data are obtained as % injected dose per gram (%ID g(-1)) from each organ, and the results correlate well with the corresponding microPET images, revealing that IV-injected mice have significantly higher uptake (p < 0.05) in the liver (5 h = 8.96% ID g(-1); 24 h = 8.27% ID g(-1)) than IR-injected mice (5 h = 0.09% ID g(-1); 24 h = 0.08% ID g(-1)). IR-injected mice show localized uptake in the large intestine (5 h = 10.37% ID g(-1); 24 h = 0.42% ID g(-1)) with minimal uptake in other organs. Raman imaging of excised tissues correlate well with biodistribution data. These results suggest that the topical application of SERS nanoparticles in the mouse colon appears to minimize their systemic distribution, thus avoiding potential toxicity and supporting the clinical translation of Raman spectroscopy as an endoscopic imaging tool.

  View details for DOI 10.1002/smll.201002317

  View details for Web of Science ID 000294361200015

  View details for PubMedID 21608124

• Bioluminescence resonance energy transfer (BRET) imaging of protein-protein interactions within deep tissues of living subjects PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Dragulescu-Andrasi, A., Chan, C. T., De, A., Massoud, T. F., Gambhir, S. S. 2011; 108 (29): 12060-12065

  Abstract

  Identifying protein-protein interactions (PPIs) is essential for understanding various disease mechanisms and developing new therapeutic approaches. Current methods for assaying cellular intermolecular interactions are mainly used for cells in culture and have limited use for the noninvasive assessment of small animal disease models. Here, we describe red light-emitting reporter systems based on bioluminescence resonance energy transfer (BRET) that allow for assaying PPIs both in cell culture and deep tissues of small animals. These BRET systems consist of the recently developed Renilla reniformis luciferase (RLuc) variants RLuc8 and RLuc8.6, used as BRET donors, combined with two red fluorescent proteins, TagRFP and TurboFP635, as BRET acceptors. In addition to the native coelenterazine luciferase substrate, we used the synthetic derivative coelenterazine-v, which further red-shifts the emission maxima of Renilla luciferases by 35 nm. We show the use of these BRET systems for ratiometric imaging of both cells in culture and deep-tissue small animal tumor models and validate their applicability for studying PPIs in mice in the context of rapamycin-induced FK506 binding protein 12 (FKBP12)-FKBP12 rapamycin binding domain (FRB) association. These red light-emitting BRET systems have great potential for investigating PPIs in the context of drug screening and target validation applications.

  View details for DOI 10.1073/pnas.1100923108

  View details for PubMedID 21730157

• Pilot Pharmacokinetic and Dosimetric Studies of F-18-FPPRGD2: A PET Radiopharmaceutical Agent for Imaging alpha(v)beta(3) Integrin Levels RADIOLOGY Mittra, E. S., Goris, M. L., Iagaru, A. H., Kardan, A., Burton, L., Berganos, R., Chang, E., Liu, S., Shen, B., Chin, F. T., Chen, X., Gambhir, S. S. 2011; 260 (1): 182-191

  Abstract

  To assess the safety, biodistribution, and dosimetric properties of the positron emission tomography (PET) radiopharmaceutical agent fluorine 18 ((18)F) FPPRGD2 (2-fluoropropionyl labeled PEGylated dimeric RGD peptide [PEG3-E{c(RGDyk)}2]), which is based on the dimeric arginine-glycine-aspartic acid (RGD) peptide sequence and targets α(v)β(3) integrin, in the first volunteers imaged with this tracer.The protocol was approved by the institutional review board, and written informed consent was obtained from all participants. Five healthy volunteers underwent whole-body combined PET-computed tomography 0.5, 1.0, 2.0, and 3.0 hours after tracer injection (mean dose, 9.5 mCi ± 3.4 [standard deviation] [351.5 MBq ± 125.8]; mean specific radioactivity, 1200 mCi/mmol ± 714 [44.4 GBq/mmol ± 26.4]). During this time, standard vital signs, electrocardiographic (ECG) readings, and blood sample values (for chemistry, hematologic, and liver function tests) were checked at regular intervals and 1 and 7 days after the injection. These data were used to evaluate tracer biodistribution and dosimetric properties, time-activity curves, and the stability of laboratory values. Significant changes in vital signs and laboratory values were evaluated by using a combination of population-averaged generalized estimating equation regression and exact paired Wilcoxon tests.The administration of (18)F-FPPRGD2 was well tolerated, with no marked effects on vital signs, ECG readings, or laboratory values. The tracer showed the same pattern of biodistribution in all volunteers: primary clearance through the kidneys (0.360 rem/mCi ± 0.185 [0.098 mSv/MBq ± 0.050]) and bladder (0.862 rem/mCi ± 0.436 [0.233 mSv/MBq ± 0.118], voiding model) and uptake in the spleen (0.250 rem/mCi ± 0.168 [0.068 mSv/MBq ± 0.046]) and large intestine (0.529 rem/mCi ± 0.236 [0.143 mSv/MBq ± 0.064]). The mean effective dose of (18)F-FPPRGD2 was 0.1462 rem/mCi ± 0.0669 (0.0396 mSv/MBq ± 0.0181). With an injected dose of 10 mCi (370 MBq) and a 1-hour voiding interval, a patient would be exposed to an effective radiation dose of 1.5 rem (15 mSv). Above the diaphragm, there was minimal uptake in the brain ventricles, salivary glands, and thyroid gland. Time-activity curves showed rapid clearance from the vasculature, with a mean 26% ± 17 of the tracer remaining in the circulation at 30 minutes and most of the activity occurring in the plasma relative to cells (mean whole blood-plasma ratio, 0.799 ± 0.096).(18)F-FPPRGD2 has desirable pharmacokinetic and biodistribution properties. The primary application is likely to be PET evaluation of oncologic patients-especially those with brain, breast, or lung cancer. Specific indications may include tumor staging, identifying patients who would benefit from antiangiogenesis therapy, and separating treatment responders from nonresponders early.

  View details for DOI 10.1148/radiol.11101139

  View details for Web of Science ID 000291932300021

  View details for PubMedID 21502381

  View details for PubMedCentralID PMC3121013

• Synthesis and Radioluminescence of PEGylated Eu3+-doped Nanophosphors as Bioimaging Probes ADVANCED MATERIALS Sun, C., Pratx, G., Carpenter, C. M., Liu, H., Cheng, Z., Gambhir, S. S., Xing, L. 2011; 23 (24): H195-H199

  View details for DOI 10.1002/adma.201100919

  View details for Web of Science ID 000293046600018

  View details for PubMedID 21557339

  View details for PubMedCentralID PMC4145869

• Therapeutic treatment of critical limb ischemia using human induced pluripotent stem cell-derived endothelial cells Huang, N. F., Jalil, R. A., Lee, J., Jame, S., Nguyen, H. N., De, A., Gambhir, S., Reijo-Pera, R., Cooke, J. P. SAGE PUBLICATIONS LTD. 2011: 221–21

  View details for Web of Science ID 000291218600013

• Potent, tumor-specific gene expression in an orthotopic hepatoma rat model using a Survivin-targeted, amplifiable adenoviral vector GENE THERAPY Ahn, B., Ronald, J. A., Kim, Y. I., Katzenberg, R., Singh, A., Paulmurugan, R., Ray, S., Hofmann, L. V., Gambhir, S. S. 2011; 18 (6): 606-612

  Abstract

  Ideal cancer gene therapies should have high tumor specificity and efficacy, and allow systemic administration to target metastases. We recently developed a bi-directional, two-step transcriptional amplification (TSTA) system driven by the tumor-specific Survivin promoter (pSurv) to amplify the correlated expression of both the reporter gene firefly luciferase (FL) and therapeutic gene tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Here, we compare the specificity and potency of an adenovirus carrying this system (Ad-pSurv-TSTA-TRAIL-FL) to a nonspecific vector (Ad-pCMV-FL) in an orthotopic hepatocellular carcinoma (HCC) rat model after systemic administration. At 24 h after injection of Ad-pCMV-FL, bioluminescence imaging revealed a trend (P=0.30) towards greater FL expression in liver versus tumor. In striking contrast, Ad-pSurv-TSTA-TRAIL-FL showed increased FL activity within the tumor compared with the liver (P<0.01), a strong trend towards reduced liver expression compared with Ad-pCMV-FL (P=0.07), and importantly, similar FL levels within tumor compared with Ad-pCMV-FL (P=0.32). Hence, this vector shows potent, tumor-specific transgene expression even after extensive liver transduction and may be of significant value in avoiding hepatotoxicity in HCC patients. Future studies will explore the benefits of tumor-specific TRAIL expression in this model, the potential to target metastases and the extension of this vector for the treatment of other Survivin-positive tumors is warranted.

  View details for DOI 10.1038/gt.2011.5

  View details for Web of Science ID 000291438900010

  View details for PubMedID 21307888

  View details for PubMedCentralID PMC4154811

• Nanoparticle PEGylation for imaging and therapy NANOMEDICINE Jokerst, J. V., Lobovkina, T., Zare, R. N., Gambhir, S. S. 2011; 6 (4): 715-728

  Abstract

  Nanoparticles are an essential component in the emerging field of nanomedical imaging and therapy. When deployed in vivo, these materials are typically protected from the immune system by polyethylene glycol (PEG). A wide variety of strategies to coat and characterize nanoparticles with PEG has established important trends on PEG size, shape, density, loading level, molecular weight, charge and purification. Strategies to incorporate targeting ligands are also prevalent. This article presents a background to investigators new to stealth nanoparticles, and suggests some key considerations needed prior to designing a nanoparticle PEGylation protocol and characterizing the performance features of the product.

  View details for DOI 10.2217/NNM.11.19

  View details for Web of Science ID 000292994300019

  View details for PubMedID 21718180

  View details for PubMedCentralID PMC3217316

• Engineering and Visualization of Bacteria for Targeting Infarcted Myocardium MOLECULAR THERAPY Le, U. N., Kim, H., Kwon, J., Kim, M. Y., Nguyen, V. H., Jiang, S. N., Lee, B., Hong, Y., Shin, M. G., Rhee, J. H., Bom, H., Ahn, Y., Gambhir, S. S., Choy, H. E., Min, J. 2011; 19 (5): 951-959

  Abstract

  Optimization of the specific affinity of cardiac delivery vector could significantly improve the efficiency of gene/protein delivery, yet no cardiac vectors to date have sufficient target specificity for myocardial infarction (MI). In this study, we explored bacterial tropism for infarcted myocardium based on our previous observations that certain bacteria are capable of targeting the hypoxic regions in solid tumors. Out of several Escherichia coli or Salmonella typhimurium strains, the S. typhimurium defective in the synthesis of ppGpp (ΔppGpp S. typhimurium) revealed accumulation and selective proliferation in the infarcted myocardium without spillover to noncardiac tissue. The Salmonellae that were engineered to express a variant of Renilla luciferase gene (RLuc8), under the control of the E. coli arabinose operon promoter (P(BAD)), selectively targeted and delivered RLuc8 in the infarcted myocardium only upon injection of L-arabinose. An examination of the infarct size before and after infection, and estimations of C-reactive protein (CRP) and procalcitonin indicated that intravenous injection of ΔppGpp S. typhimurium did not induce serious local or systemic immune reactions. This current proof-of-principle study demonstrates for the first time the capacity of Salmonellae to target infarcted myocardium and to serve as a vehicle for the selective delivery of therapeutic agents in MI.

  View details for DOI 10.1038/mt.2011.25

  View details for Web of Science ID 000290146200021

  View details for PubMedID 21364539

  View details for PubMedCentralID PMC3098638

• Combined 18F NaF and 18F FDG PET/CT: Initial results of a multi-center trial Iagaru, A., Mittra, E., Sathekge, M., Prakash, V., Iyer, V., Dick, D., Lapa, P., Isidoro, J., de Lima, J., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2011

  View details for Web of Science ID 000443798900035

• 18F FPPRGD2 in breast cancer subjects: A novel PET radiopharmaceutical for imaging alpha v beta 3 integrin levels Iagaru, A., Mosci, C., Mittra, E., Shen, B., Chin, F., Chen, X., Telli, M., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2011

  View details for Web of Science ID 000443798900075

• Studies of the 18F L-glutamate derivative BAY 94-9392 in cancer patients: A novel radiopharmaceutical for PET imaging Mittra, E., Mosci, C., Iagaru, A., Fels, L., Bacher-Stier, C., Chin, F., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2011

  View details for Web of Science ID 000443798902382

• Prediction of human PET imaging dose to monitor NHL therapy using < 64 > Cu-DOTA-rituximab and a transgenic mouse model Natarajan, A., Gowrishankar, G., Nielsen, C., Wang, S., Iagaru, A., Goris, M., Gambhir, S., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2011

  View details for Web of Science ID 000443798902179

• Exploring Gaussia luciferase as a novel secretion system for eukaryotic protein delivery for bacteria-mediated cancer treatment Vu Nguyen, Thuy Phan, Paulmurugan, R., Gambhir, S., Min, J. SOC NUCLEAR MEDICINE INC. 2011

  View details for Web of Science ID 000443798902153

• Multimodality Imaging of beta-Cells in Mouse Models of Type 1 and 2 Diabetes DIABETES Yong, J., Rasooly, J., Dang, H., Lu, Y., Middleton, B., Zhang, Z., Hon, L., Namavari, M., Stout, D. B., Atkinson, M. A., Tian, J., Gambhir, S. S., Kaufman, D. L. 2011; 60 (5): 1383-1392

  Abstract

  β-Cells that express an imaging reporter have provided powerful tools for studying β-cell development, islet transplantation, and β-cell autoimmunity. To further expedite diabetes research, we generated transgenic C57BL/6 "MIP-TF" mice that have a mouse insulin promoter (MIP) driving the expression of a trifusion (TF) protein of three imaging reporters (luciferase/enhanced green fluorescent protein/HSV1-sr39 thymidine kinase) in their β-cells. This should enable the noninvasive imaging of β-cells by charge-coupled device (CCD) and micro-positron emission tomography (PET), as well as the identification of β-cells at the cellular level by fluorescent microscopy.MIP-TF mouse β-cells were multimodality imaged in models of type 1 and type 2 diabetes.MIP-TF mouse β-cells were readily identified in pancreatic tissue sections using fluorescent microscopy. We show that MIP-TF β-cells can be noninvasively imaged using microPET. There was a correlation between CCD and microPET signals from the pancreas region of individual mice. After low-dose streptozotocin administration to induce type 1 diabetes, we observed a progressive reduction in bioluminescence from the pancreas region before the appearance of hyperglycemia. Although there have been reports of hyperglycemia inducing proinsulin expression in extrapancreatic tissues, we did not observe bioluminescent signals from extrapancreatic tissues of diabetic MIP-TF mice. Because MIP-TF mouse β-cells express a viral thymidine kinase, ganciclovir treatment induced hyperglycemia, providing a new experimental model of type 1 diabetes. Mice fed a high-fat diet to model early type 2 diabetes displayed a progressive increase in their pancreatic bioluminescent signals, which were positively correlated with area under the curve-intraperitoneal glucose tolerance test (AUC-IPGTT).MIP-TF mice provide a new tool for monitoring β-cells from the single cell level to noninvasive assessments of β-cells in models of type 1 diabetes and type 2 diabetes.

  View details for DOI 10.2337/db10-0907

  View details for Web of Science ID 000290349700004

  View details for PubMedID 21441442

  View details for PubMedCentralID PMC3292311

• Early Diagnosis of Ovarian Carcinoma: Is a Solution in Sight? RADIOLOGY Lutz, A. M., Willmann, J. K., Drescher, C. W., Ray, P., Cochran, F. V., Urban, N., Gambhir, S. S. 2011; 259 (2): 329-345

  Abstract

  Ovarian cancer is the most lethal of the gynecologic malignancies. Because ovarian cancer symptoms are subtle and nonspecific, the diagnosis is often delayed until the disease is well advanced. Overall 5-year survival is a rather dismal 50% but can be improved to greater than 90% if the disease is confined to the ovary at the time of diagnosis (generally in fewer than 25% of patients). Effective screening tools are currently not available. Owing to the rather low incidence of the disease in the general population, potential screening tests must provide very high specificity to avoid unnecessary interventions in false-positive cases. This article reviews currently available serum biomarkers and imaging tests for the early detection of ovarian cancer and provides an outlook on the potential improvements in these noninvasive diagnostic tools that may lead to successful implementation in a screening program. Supplemental material: http://radiology.rsna.org/lookup/suppl/doi:10.1148/radiol.11090563/-/DC1.

  View details for DOI 10.1148/radiol.11090563

  View details for Web of Science ID 000289667300006

  View details for PubMedID 21502390

• "Same Day" Ex-vivo Regional Gene Therapy: A Novel Strategy to Enhance Bone Repair MOLECULAR THERAPY Virk, M. S., Sugiyama, O., Park, S. H., Gambhir, S. S., Adams, D. J., Drissi, H., Lieberman, J. R. 2011; 19 (5): 960-968

  Abstract

  Ex-vivo regional gene therapy with bone marrow cells (BMCs) overexpressing bone morphogenetic protein-2 (BMP-2) has demonstrated efficacy in healing critical sized bone defects in preclinical studies. The purpose of this preclinical study was to compare the osteoinductive potential of a novel "same day" ex-vivo regional gene therapy versus a traditional two-step approach, which involves culture expansion of the donor cells before implantation. In the "same day" strategy buffy coat cells were harvested from the rat bone marrow, transduced with a lentiviral vector-expressing BMP-2 for 1 hour and implanted into a rat femoral defect in the same sitting. There was no significant difference (P = 0.22) with respect to the radiographic healing rates between the femoral defects treated with the "same day" strategy (13/13; 100%) versus the traditional two-step approach (11/14; 78%). However, the femoral defects treated with the "same day" strategy induced earlier radiographic bone healing (P = 0.004) and higher bone volume (BV) [micro-computed tomography (micro-CT); P < 0.001]. The "same day" strategy represents a significant advance in the field of ex-vivo regional gene therapy because it offers a solution to limitations associated with the culture expansion process required in the traditional ex vivo approach. This strategy should be cost-effective when adapted for human use.

  View details for DOI 10.1038/mt.2011.2

  View details for Web of Science ID 000290146200022

  View details for PubMedID 21343916

  View details for PubMedCentralID PMC3098640

• The Fate and Toxicity of Raman-Active Silica-Gold Nanoparticles in Mice SCIENCE TRANSLATIONAL MEDICINE Thakor, A. S., Luong, R., Paulmurugan, R., Lin, F. I., Kempen, P., Zavaleta, C., Chu, P., Massoud, T. F., Sinclair, R., Gambhir, S. S. 2011; 3 (79)

  Abstract

  Raman spectroscopy is an optical imaging method that is based on the Raman effect, the inelastic scattering of a photon when energy is absorbed from light by a surface. Although Raman spectroscopy is widely used for chemical and molecular analysis, its clinical application has been hindered by the inherently weak nature of the Raman effect. Raman-silica-gold-nanoparticles (R-Si-Au-NPs) overcome this limitation by producing larger Raman signals through surface-enhanced Raman scattering. Because we are developing these particles for use as targeted molecular imaging agents, we examined the acute toxicity and biodistribution of core polyethylene glycol (PEG)-ylated R-Si-Au-NPs after different routes of administration in mice. After intravenous administration, PEG-R-Si-Au-NPs were removed from the circulation by macrophages in the liver and spleen (that is, the reticuloendothelial system). At 24 hours, PEG-R-Si-Au-NPs elicited a mild inflammatory response and an increase in oxidative stress in the liver, which subsided by 2 weeks after administration. No evidence of significant toxicity was observed by measuring clinical, histological, biochemical, or cardiovascular parameters for 2 weeks. Because we are designing targeted PEG-R-Si-Au-NPs (for example, PEG-R-Si-Au-NPs labeled with an affibody that binds specifically to the epidermal growth factor receptor) to detect colorectal cancer after administration into the bowel lumen, we tested the toxicity of the core nanoparticle after administration per rectum. We observed no significant bowel or systemic toxicity, and no PEG-R-Si-Au-NPs were detected systemically. Although additional studies are required to investigate the long-term effects of PEG-R-Si-Au-NPs and their toxicity when carrying the targeting moiety, the results presented here support the idea that PEG-R-Si-Au-NPs can be safely used in living subjects, especially when administered rectally.

  View details for DOI 10.1126/scitranslmed.3001963

  View details for PubMedID 21508310

• F-18-FAZA-PET as a pharmacodynamic biomarker of anti-tumor activity for the novel HIF-1 alpha pathway inhibitor BAY 87-2243 in preclinical tumor models Chan, E., Liu, S., Berhoerster, K., Ellinghaus, P., Liu, S., Gekeler, V., Cheng, Z., Berndorff, D., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2011

  View details for DOI 10.1158/1538-7445.AM2011-LB-147

  View details for Web of Science ID 000209701305466

• A novel method of tumor characterization by protein and microRNA biomarker release using ultrasound D'Souza, A. L., Chevillet, J. R., Kroh, E. M., Tewari, M., Gambhir, S. S., Glazer, G. M. AMER ASSOC CANCER RESEARCH. 2011

  View details for DOI 10.1158/1538-7445.AM2011-4881

  View details for Web of Science ID 000209701305199

• A mathematical approach to earlier cancer detection: Using blood biomarker assays to monitor growth of a tumor from a single cell Hori, S. S., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2011

  View details for DOI 10.1158/1538-7445.AM2011-5079

  View details for Web of Science ID 000209701405059

• FDG-PET/CT in Cancers of the Head and Neck: What is the Definition of Whole Body Scanning? MOLECULAR IMAGING AND BIOLOGY Iagaru, A., Mittra, E. S., Gambhir, S. S. 2011; 13 (2): 362-367

  Abstract

  The role of 2-deoxy-2-[F-18]fluoro-D-glucose-positron emission tomography (FDG-PET) was studied in a variety of cancers, including head and neck squamous cell carcinomas (HNSCC) and nasopharyngeal carcinomas (NPC), with several presentations indicating that for these clinical entities a "whole-body" (i.e., eyes to thighs) may yield little additional information. Therefore, we were prompted to review our experience with PET/computed tomography (CT) in the management of patients with HNSCC and NPC.This is a retrospective study of 133 patients with HNSCC, 23-90 years old (average: 58.2 ± 12.7) and 26 patients with NPC, ages 16-75 (average: 47.3 ± 17.1), who had whole body PET/CT at our institution from Jan 2003 to Nov 2006. Reinterpretation of the imaging studies for accuracy and data analysis from medical records was performed. Lesions identified on PET/CT below the level of the adrenal glands were recorded and tabulated.Lesions were identified below the adrenal glands in seven patients (5.2%) with HNSCC. These included hepatic and osseous metastases from HNSCC in two patients (1.5%), a new renal cancer (0.75%), a new pancreatic cancer (0.75%), a new colon cancer (0.75%) and findings proven benign on follow-up (focal colon uptake in one patient and an inflammatory inguinal lymph node in another patient; 1.5%). Lesions were identified below the adrenal glands in three patients (11.5%) with NPC. These included osseous metastases from NPC in two patients (7.7%) and findings proven benign on follow-up (focal colon uptake in one patient; 3.84%).This study suggests that whole body PET/CT imaging in HNSCC has a relatively low yield (3%, 95% CI: 1.33-8.42) of significant findings below the level of the adrenal glands. Therefore, implementing a more limited protocol (through the level of adrenal glands), especially in low-risk cases of HNSCC, may be considered. However, whole body PET/CT imaging in NPC may have a significant yield (7.7%, 95% CI: 1.02-25.26) of medically relevant findings below the level of the adrenal glands. Thus, the whole body (i.e., vertex to thighs) PET/CT scan of NPC patients appears to be the appropriate imaging protocol for this population. This recommendation requires further evaluation and validation in larger prospective studies.

  View details for DOI 10.1007/s11307-010-0343-8

  View details for Web of Science ID 000288177700021

  View details for PubMedID 20495879

• Reproducibility study of [F-18]FPP(RGD)(2) uptake in murine models of human tumor xenografts EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Chang, E., Liu, S., Gowrishankar, G., Yaghoubi, S., Wedgeworth, J. P., Chin, F., Berndorff, D., Gekeler, V., Gambhir, S. S., Cheng, Z. 2011; 38 (4): 722-730

  Abstract

  An (18)F-labeled PEGylated arginine-glycine-aspartic acid (RGD) dimer {[(18)F]FPP(RGD)(2)} has been used to image tumor α(v)β(3) integrin levels in preclinical and clinical studies. Serial positron emission tomography (PET) studies may be useful for monitoring antiangiogenic therapy response or for drug screening; however, the reproducibility of serial scans has not been determined for this PET probe. The purpose of this study was to determine the reproducibility of the integrin α(v)β(3)-targeted PET probe, [(18)F]FPP(RGD)(2,) using small animal PET.Human HCT116 colon cancer xenografts were implanted into nude mice (n = 12) in the breast and scapular region and grown to mean diameters of 5-15 mm for approximately 2.5 weeks. A 3-min acquisition was performed on a small animal PET scanner approximately 1 h after administration of [(18)F]FPP(RGD)(2) (1.9-3.8 MBq, 50-100 μCi) via the tail vein. A second small animal PET scan was performed approximately 6 h later after reinjection of the probe to assess for reproducibility. Images were analyzed by drawing an ellipsoidal region of interest (ROI) around the tumor xenograft activity. Percentage injected dose per gram (%ID/g) values were calculated from the mean or maximum activity in the ROIs. Coefficients of variation and differences in %ID/g values between studies from the same day were calculated to determine the reproducibility.The coefficient of variation (mean±SD) for %ID(mean)/g and %ID(max)/g values between [(18)F]FPP(RGD)(2) small animal PET scans performed 6 h apart on the same day were 11.1 ± 7.6% and 10.4 ± 9.3%, respectively. The corresponding differences in %ID(mean)/g and %ID(max)/g values between scans were -0.025 ± 0.067 and -0.039 ± 0.426. Immunofluorescence studies revealed a direct relationship between extent of α(ν)β(3) integrin expression in tumors and tumor vasculature with level of tracer uptake. Mouse body weight, injected dose, and fasting state did not contribute to the variability of the scans; however, consistent scanning parameters were necessary to ensure accurate studies, in particular, noting tumor volume, as well as making uniform: the time of imaging after injection and the ROI size. Reanalysis of ROI placement displayed variability for %ID(mean)/g of 6.6 ± 3.9% and 0.28 ± 0.12% for %ID(max)/g.[(18)F]FPP(RGD)(2) small animal PET mouse tumor xenograft studies are reproducible with relatively low variability.

  View details for DOI 10.1007/s00259-010-1672-1

  View details for Web of Science ID 000288255500015

  View details for PubMedID 21125268

• Molecular Imaging Using Light-Absorbing Imaging Agents and a Clinical Optical Breast Imaging System-a Phantom Study MOLECULAR IMAGING AND BIOLOGY van de Ven, S. M., Mincu, N., Brunette, J., Ma, G., Khayat, M., Ikeda, D. M., Gambhir, S. S. 2011; 13 (2): 232-238

  Abstract

  The aim of the study was to determine the feasibility of using a clinical optical breast scanner with molecular imaging strategies based on modulating light transmission.Different concentrations of single-walled carbon nanotubes (SWNT; 0.8-20.0 nM) and black hole quencher-3 (BHQ-3; 2.0-32.0 µM) were studied in specifically designed phantoms (200-1,570 mm(3)) with a clinical optical breast scanner using four wavelengths. Each phantom was placed in the scanner tank filled with optical matching medium. Background scans were compared to absorption scans, and reproducibility was assessed.All SWNT phantoms were detected at four wavelengths, with best results at 684 nm. Higher concentrations (≥8.0 µM) were needed for BHQ-3 detection, with the largest contrast at 684 nm. The optical absorption signal was dependent on phantom size and concentration. Reproducibility was excellent (intraclass correlation 0.93-0.98).Nanomolar concentrations of SWNT and micromolar concentrations of BHQ-3 in phantoms were reproducibly detected, showing the potential of light absorbers, with appropriate targeting ligands, as molecular imaging agents for clinical optical breast imaging.

  View details for DOI 10.1007/s11307-010-0356-3

  View details for Web of Science ID 000288177700006

  View details for PubMedID 20532642

• Multiparametric MRI reveals early response patterns to antiangiogenic therapy in primary breast cancer HUGHES, N. P., Mehta, S., Adams, R. F., Gambhir, S. S., Padhani, A. R., Harris, A. L. SOC NUCLEAR MEDICINE INC. 2011: 668–69

  View details for Web of Science ID 000288804500051

• Molecular imaging of the Epidermal Growth Factor Receptor in rodent colon via Affibody-functionalized surface enhanced Raman scattering (SERS) nanoparticles 241st National Meeting and Exposition of the American-Chemical-Society (ACS) Jokerst, J. V., Miao, Z., Thakor, A. S., Cheng, Z., Gambhir, S. S. AMER CHEMICAL SOC. 2011

  View details for Web of Science ID 000291982802272

• MYC Phosphorylation, Activation, and Tumorigenic Potential in Hepatocellular Carcinoma Are Regulated by HMG-CoA Reductase CANCER RESEARCH Cao, Z., Fan-Minogue, H., Bellovin, D. I., Yevtodiyenko, A., Arzeno, J., Yang, Q., Gambhir, S. S., Felsher, D. W. 2011; 71 (6): 2286-2297

  Abstract

  MYC is a potential target for many cancers but is not amenable to existing pharmacologic approaches. Inhibition of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMG-CoA reductase) by statins has shown potential efficacy against a number of cancers. Here, we show that inhibition of HMG-CoA reductase by atorvastatin (AT) blocks both MYC phosphorylation and activation, suppressing tumor initiation and growth in vivo in a transgenic model of MYC-induced hepatocellular carcinoma (HCC) as well as in human HCC-derived cell lines. To confirm specificity, we show that the antitumor effects of AT are blocked by cotreatment with the HMG-CoA reductase product mevalonate. Moreover, by using a novel molecular imaging sensor, we confirm that inhibition of HMG-CoA reductase blocks MYC phosphorylation in vivo. Importantly, the introduction of phosphorylation mutants of MYC at Ser62 or Thr58 into tumors blocks their sensitivity to inhibition of HMG-CoA reductase. Finally, we show that inhibition of HMG-CoA reductase suppresses MYC phosphorylation through Rac GTPase. Therefore, HMG-CoA reductase is a critical regulator of MYC phosphorylation, activation, and tumorigenic properties. The inhibition of HMG-CoA reductase may be a useful target for the treatment of MYC-associated HCC as well as other tumors.

  View details for DOI 10.1158/0008-5472.CAN-10-3367

  View details for Web of Science ID 000288381300028

  View details for PubMedID 21262914

  View details for PubMedCentralID PMC3059327

• Affibody-Functionalized Gold-Silica Nanoparticles for Raman Molecular Imaging of the Epidermal Growth Factor Receptor SMALL Jokerst, J. V., Miao, Z., Zavaleta, C., Cheng, Z., Gambhir, S. S. 2011; 7 (5): 625-633

  Abstract

  The affibody functionalization of fluorescent surface-enhanced Raman scattering gold-silica nanoparticles as multimodal contrast agents for molecular imaging specific to epidermal growth factor receptor (EGFR) is reported. This nanoparticle bioconjugate reports EGFR-positive A431 tumors with a signal nearly 35-fold higher than EGFR-negative MDA-435S tumors. The low-level EGFR expression in adjacent healthy tissue is 7-fold lower than in the positive tumors. Validation via competitive inhibition reduces the signal by a factor of six, and independent measurement of EGFR via flow cytometry correlates at R(2) = 0.92.

  View details for DOI 10.1002/smll.201002291

  View details for Web of Science ID 000288081900013

  View details for PubMedID 21302357

  View details for PubMedCentralID PMC3386295

• Affibody-based nanoprobes for HER2-expressing cell and tumor imaging BIOMATERIALS Gao, J., Chen, K., Miao, Z., Ren, G., Chen, X., Gambhir, S. S., Cheng, Z. 2011; 32 (8): 2141-2148

  Abstract

  This article reports the affibody-based nanoprobes specifically target and image human epidermal growth factor receptor type 2 (HER2)-expressing cells and tumors. The affibody molecules are a promising class of targeting ligands with simple, robust, and precise structure and high affinity. Using near-infrared (NIR) quantum dots (QDs) and iron oxide (IO) nanoparticles as two representative nanomaterials, we designed anti-HER2 affibody molecules with a N-terminus cysteine residue (Cysteine-Z(HER2:342)) and precisely conjugated with maleimide-functionalized nanoparticles to make nanoparticle-affibody conjugates. The in vitro and in vivo study showed the conjugates are highly specific to target and image HER2-expressing cells and tumors. This work indicated the nanoparticle-affibody conjugates may be excellent candidates as targeting probes for molecular imaging and diagnosis.

  View details for DOI 10.1016/j.biomaterials.2010.11.053

  View details for Web of Science ID 000287061400015

  View details for PubMedID 21147502

  View details for PubMedCentralID PMC3032351

• Use of DNA Microarray and Small Animal Positron Emission Tomography in Preclinical Drug Evaluation of RAF265, a Novel B-Raf/VEGFR-2 lInhibitor NEOPLASIA Tseng, J. R., Stuart, D., Aardalen, K., Kaplan, A., Aziz, N., Hughes, N. P., Gambhir, S. S. 2011; 13 (3): 266-U108

  Abstract

  Positron emission tomography (PET) imaging has become a useful tool for assessing early biologic response to cancer therapy and may be particularly useful in the development of new cancer therapeutics. RAF265, a novel B-Raf/vascular endothelial growth factor receptor-2 inhibitor, was evaluated in the preclinical setting for its ability to inhibit the uptake of PET tracers in the A375M(B-Raf(V600E)) human melanoma cell line. RAF265 inhibited 2-deoxy-2-[(18)F]fluoro-d-glucose (FDG) accumulation in cell culture at 28 hours in a dose-dependent manner. RAF265 also inhibited FDG accumulation in tumor xenografts after 1 day of drug treatment. This decrease persisted for the remaining 2 weeks of treatment. DNA microarray analysis of treated tumor xenografts revealed significantly decreased expression of genes regulating glucose and thymidine metabolism and revealed changes in apoptotic genes, suggesting that the imaging tracers FDG, 3-deoxy-3-[(18)F]fluorothymidine, and annexin V could serve as potential imaging biomarkers for RAF265 therapy monitoring. We concluded that RAF265 is highly efficacious in this xenograft model of human melanoma and decreases glucose metabolism as measured by DNA microarray analysis, cell culture assays, and small animal FDG PET scans as early as 1 day after treatment. Our results support the use of FDG PET in clinical trials with RAF265 to assess early tumor response. DNA microarray analysis and small animal PET studies may be used as complementary technologies in drug development. DNA microarray analysis allows for analysis of drug effects on multiple pathways linked to cancer and can suggest corresponding imaging tracers for further analysis as biomarkers of tumor response.

  View details for DOI 10.1593/neo.101466

  View details for PubMedID 21390189

• Noninvasive Monitoring of Placenta-Specific Transgene Expression by Bioluminescence Imaging PLOS ONE Fan, X., Ren, P., Dhal, S., Bejerano, G., Goodman, S. B., Druzin, M. L., Gambhir, S. S., Nayak, N. R. 2011; 6 (1)

  Abstract

  Placental dysfunction underlies numerous complications of pregnancy. A major obstacle to understanding the roles of potential mediators of placental pathology has been the absence of suitable methods for tissue-specific gene manipulation and sensitive assays for studying gene functions in the placentas of intact animals. We describe a sensitive and noninvasive method of repetitively tracking placenta-specific gene expression throughout pregnancy using lentivirus-mediated transduction of optical reporter genes in mouse blastocysts.Zona-free blastocysts were incubated with lentivirus expressing firefly luciferase (Fluc) and Tomato fluorescent fusion protein for trophectoderm-specific infection and transplanted into day 3 pseudopregnant recipients (GD3). Animals were examined for Fluc expression by live bioluminescence imaging (BLI) at different points during pregnancy, and the placentas were examined for tomato expression in different cell types on GD18. In another set of experiments, blastocysts with maximum photon fluxes in the range of 2.0E+4 to 6.0E+4 p/s/cm(2)/sr were transferred. Fluc expression was detectable in all surrogate dams by day 5 of pregnancy by live imaging, and the signal increased dramatically thereafter each day until GD12, reaching a peak at GD16 and maintaining that level through GD18. All of the placentas, but none of the fetuses, analyzed on GD18 by BLI showed different degrees of Fluc expression. However, only placentas of dams transferred with selected blastocysts showed uniform photon distribution with no significant variability of photon intensity among placentas of the same litter. Tomato expression in the placentas was limited to only trophoblast cell lineages.These results, for the first time, demonstrate the feasibility of selecting lentivirally-transduced blastocysts for uniform gene expression in all placentas of the same litter and early detection and quantitative analysis of gene expression throughout pregnancy by live BLI. This method may be useful for a wide range of applications involving trophoblast-specific gene manipulations in utero.

  View details for DOI 10.1371/journal.pone.0016348

  View details for PubMedID 21283713

• Oxidative Stress Mediates the Effects of Raman-Active Gold Nanoparticles in Human Cells SMALL Thakor, A. S., Paulmurugan, R., Kempen, P., Zavaleta, C., Sinclair, R., Massoud, T. F., Gambhir, S. S. 2011; 7 (1): 126-136

  Abstract

  Polyethylene glycol (PEG)ylated Raman-active gold nanoparticles (PEG-R-AuNPs) consist of an interchangeable Raman organic molecule layer held onto a gold nanocore by a silica shell. PEG-R-AuNPs have been shown preclinically to increase the sensitivity and specificity of Raman spectroscopy, with picomolar sensitivity and multiplexing capabilities. Although clinical trials are being designed to use functionalized PEG-R-AuNPs in various applications (e.g., to target dysplastic bowel lesions during colonoscopy), the effects of these nanoparticles on human cells remain unknown. The occurrence and mechanisms underlying any potential cytotoxicity induced by these nanoparticles (0-1000 PEG-R-AuNPs/cell) are investigated in immortalized human HeLa and HepG2 cell lines at several time points (0-48 h) after exposure. Using fluorometric assays, cell viability (MTT), reactive oxygen species (ROS) generation (dichlorofluorescein diacetate), protein oxidation (protein carbonyl content), and total cellular antioxidant concentrations the concentrations (metmyoblobin-induced oxidation of ABTS) are assessed. Analysis of lipid oxidation using an enzyme immunoassay (8-isoprostane concentrations), gene expression of antioxidant enzymes using quantitative reverse transcription polymerase chain reactions, and the intracellular location of PEG-R-AuNPs using transmission electron microscopy is also undertaken. PEG-R-AuNPs cause no cytotoxicity in either HeLa or HepG2 cells in the acute setting as ROS generation is balanced by antioxidant enzyme upregulation. Following prolonged exposures (48 h) at relatively high concentrations (1000 PEG-R-AuNPs/cell), nanoparticles are found within vesicles inside cells. Under these conditions, a minimal amount of cytotoxicity is seen in both cell lines owing to increases in cellular oxidative stress, most likely due to ROS overwhelming the antioxidant defenses. Evidence of oxidative stress-induced damage includes increased lipid and protein oxidation. Although further in vivo toxicity studies are necessary, these initial encouraging results show that PEG-R-AuNPs cause minimal toxicity in human cells in the acute setting, which bodes well for potential future applications of these nanoparticles in living subjects.

  View details for DOI 10.1002/smll.201001466

  View details for PubMedID 21104804

• Radiolabeling of a Saxitoxin derivative for PET-MRI imaging of pain Hoehne, A., Parsons, W. H., Behera, D., Shen, B., Gambhir, S. S., Du Bois, J., Biswal, S., Chin, F. T. WILEY-BLACKWELL. 2011: S2–S2

  View details for Web of Science ID 000295901600003

• Clinical Application of Non-Invasive Molecular Imaging Cancer Cell Therapy: The First Reporter Gene-Based Imaging Clinical Trial Yaghoubi, S., Gambhir, S. S. edited by Yaghoubi, S., Gambhir, S. S. Cambridge University Press. 2011: 369–386
• [F-18]YF3 nanoprobes: a novel synthetic strategy for F-18-labeled imaging agents Xiong Liqin, L. Q., Shen Bin, B., Gambhir, S. S., Chin, F. T., Rao Jianghong, J. H. WILEY-BLACKWELL. 2011: S77–S77

  View details for Web of Science ID 000295901600078

• [F-18]FTC-146 for imaging sigma-1 receptors in squirrel monkey brain using PET/MRI James, M. L., Shen, B., Nielsen, C. H., Buckmaster, C. L., Berganos, R. A., Zavaleta, C., Lyons, D., McCurdy, C. R., Gambhir, S. S., Chin, F. T. WILEY-BLACKWELL. 2011: S317–S317

  View details for Web of Science ID 000295901600317

• A Hybrid Least Squares and Principal Component Analysis Algorithm for Raman Spectroscopy 33rd Annual International Conference of the IEEE Engineering-in-Medicine-and-Biology-Society (EMBS) Van de Sompel, D., Garai, E., Zavaleta, C., Gambhir, S. S. IEEE. 2011: 6971–6974

  Abstract

  The least squares fitting algorithm is the most commonly used algorithm in Raman spectroscopy. In this paper, however, we show that it is sensitive to variations in the background signal when the signal of interest is weak. To address this problem, we propose a novel algorithm to analyze measured spectra in Raman spectroscopy. The method is a hybrid least squares and principal component analysis algorithm. It explicitly accounts for any variations expected in the reference spectra used in the signal decomposition. We compare the novel algorithm to the least squares method with a low-order polynomial residual model, and demonstrate the novel algorithm's superior performance by comparing quantitative error metrics. Our experiments use both simulated data and data acquired from an in vitro solution of Raman-enhanced gold nanoparticles.

  View details for Web of Science ID 000298810005123

  View details for PubMedID 22255942

• F-18-Cyanobenzolthiol ([F-18]CBT): A novel F-18-prosthetic group for labeling peptide or protein Shen Bin, B., Jeon, J., Gambhir, S. S., Rao, J., Chin, F. T. WILEY-BLACKWELL. 2011: S503–S503

  View details for Web of Science ID 000295901600503

• Longitudinal, Noninvasive Imaging of T-Cell Effector Function and Proliferation in Living Subjects CANCER RESEARCH Patel, M. R., Chang, Y., Chen, I. Y., Bachmann, M. H., Yan, X., Contag, C. H., Gambhir, S. S. 2010; 70 (24): 10141-10149

  Abstract

  Adoptive immunotherapy is evolving to assume an increasing role in treating cancer. Most imaging studies in adoptive immunotherapy to date have focused primarily on locating tumor-specific T cells rather than understanding their effector functions. In this study, we report the development of a noninvasive imaging strategy to monitor T-cell activation in living subjects by linking a reporter gene to the Granzyme B promoter (pGB), whose transcriptional activity is known to increase during T-cell activation. Because pGB is relatively weak and does not lead to sufficient reporter gene expression for noninvasive imaging, we specifically employed 2 signal amplification strategies, namely the Two Step Transcription Amplification (TSTA) strategy and the cytomegalovirus enhancer (CMVe) strategy, to maximize firefly luciferase reporter gene expression. Although both amplification strategies were capable of increasing pGB activity in activated primary murine splenocytes, only the level of bioluminescence activity achieved with the CMVe strategy was adequate for noninvasive imaging in mice. Using T cells transduced with a reporter vector containing the hybrid pGB-CMVe promoter, we were able to optically image T-cell effector function longitudinally in response to tumor antigens in living mice. This methodology has the potential to accelerate the study of adoptive immunotherapy in preclinical cancer models.

  View details for DOI 10.1158/0008-5472.CAN-10-1843

  View details for Web of Science ID 000285334200016

  View details for PubMedID 21159636

  View details for PubMedCentralID PMC3057959

• Near-infrared fluorescent nanoprobes for cancer molecular imaging: status and challenges TRENDS IN MOLECULAR MEDICINE He, X., Gao, J., Gambhir, S. S., Cheng, Z. 2010; 16 (12): 574-583

  Abstract

  Near-infrared fluorescence (NIRF) imaging promises to improve cancer imaging and management; advances in nanomaterials allow scientists to combine new nanoparticles with NIRF imaging techniques, thereby fulfilling this promise. Here, we present a synopsis of current developments in NIRF nanoprobes, their use in imaging small living subjects, their pharmacokinetics and toxicity, and finally their integration into multimodal imaging strategies. We also discuss challenges impeding the clinical translation of NIRF nanoprobes for molecular imaging of cancer. Whereas utilization of most NIRF nanoprobes remains at a proof-of-principle stage, optimizing the impact of nanomedicine in cancer patient diagnosis and management will probably be realized through persistent interdisciplinary amalgamation of diverse research fields.

  View details for DOI 10.1016/j.molmed.2010.08.006

  View details for Web of Science ID 000285727200004

  View details for PubMedID 20870460

  View details for PubMedCentralID PMC2994979

• PET Imaging of Tumor Neovascularization in a Transgenic Mouse Model with a Novel Cu-64-DOTA-Knottin Peptide CANCER RESEARCH Nielsen, C. H., Kimura, R. H., Withofs, N., Tran, P. T., Miao, Z., Cochran, J. R., Cheng, Z., Felsher, D., Kjaer, A., Willmann, J. K., Gambhir, S. S. 2010; 70 (22): 9022-9030

  Abstract

  Due to the high mortality of lung cancer, there is a critical need to develop diagnostic procedures enabling early detection of the disease while at a curable stage. Targeted molecular imaging builds on the positive attributes of positron emission tomography/computed tomography (PET/CT) to allow for a noninvasive detection and characterization of smaller lung nodules, thus increasing the chances of positive treatment outcome. In this study, we investigate the ability to characterize lung tumors that spontaneously arise in a transgenic mouse model. The tumors are first identified with small animal CT followed by characterization with the use of small animal PET with a novel 64Cu-1,4,7,10-tetra-azacylododecane-N,N',N'',N'''-tetraacetic acid (DOTA)-knottin peptide that targets integrins upregulated during angiogenesis on the tumor associated neovasculature. The imaging results obtained with the knottin peptide are compared with standard 18F-fluorodeoxyglucose (FDG) PET small animal imaging. Lung nodules as small as 3 mm in diameter were successfully identified in the transgenic mice by small animal CT, and both 64Cu-DOTA-knottin 2.5F and FDG were able to differentiate lung nodules from the surrounding tissues. Uptake and retention of the 64Cu-DOTA-knottin 2.5F tracer in the lung tumors combined with a low background in the thorax resulted in a statistically higher tumor to background (normal lung) ratio compared with FDG (6.01±0.61 versus 4.36±0.68; P<0.05). Ex vivo biodistribution showed 64Cu-DOTA-knottin 2.5F to have a fast renal clearance combined with low nonspecific accumulation in the thorax. Collectively, these results show 64Cu-DOTA-knottin 2.5F to be a promising candidate for clinical translation for earlier detection and improved characterization of lung cancer.

  View details for DOI 10.1158/0008-5472.CAN-10-1338

  View details for Web of Science ID 000284213300008

  View details for PubMedID 21062977

  View details for PubMedCentralID PMC3057960

• Cancer stem cells from human breast tumors are involved in spontaneous metastases in orthotopic mouse models PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Liu, H., Patel, M. R., Prescher, J. A., Patsialou, A., Qian, D., Lin, J., Wen, S., Chang, Y., Bachmann, M. H., Shimono, Y., Dalerba, P., Adorno, M., Lobo, N., Bueno, J., Dirbas, F. M., Goswami, S., Somlo, G., Condeelis, J., Contag, C. H., Gambhir, S. S., Clarke, M. F. 2010; 107 (42): 18115-18120

  Abstract

  To examine the role of breast cancer stem cells (BCSCs) in metastasis, we generated human-in-mouse breast cancer orthotopic models using patient tumor specimens, labeled with optical reporter fusion genes. These models recapitulate human cancer features not captured with previous models, including spontaneous metastasis in particular, and provide a useful platform for studies of breast tumor initiation and progression. With noninvasive imaging approaches, as few as 10 cells of stably labeled BCSCs could be tracked in vivo, enabling studies of early tumor growth and spontaneous metastasis. These advances in BCSC imaging revealed that CD44(+) cells from both primary tumors and lung metastases are highly enriched for tumor-initiating cells. Our metastatic cancer models, combined with noninvasive imaging techniques, constitute an integrated approach that could be applied to dissect the molecular mechanisms underlying the dissemination of metastatic CSCs (MCSCs) and to explore therapeutic strategies targeting MCSCs in general or to evaluate individual patient tumor cells and predict response to therapy.

  View details for DOI 10.1073/pnas.1006732107

  View details for Web of Science ID 000283184800050

  View details for PubMedID 20921380

  View details for PubMedCentralID PMC2964232

• Dynamic Visualization of RGD-Quantum Dot Binding to Tumor Neovasculature and Extravasation in Multiple Living Mouse Models Using Intravital Microscopy SMALL Smith, B. R., Cheng, Z., De, A., Rosenberg, J., Gambhir, S. S. 2010; 6 (20): 2222-2229

  View details for DOI 10.1002/smll.201001022

  View details for Web of Science ID 000283890500003

  View details for PubMedID 20862677

  View details for PubMedCentralID PMC3030963

• 3-D Deep Penetration Photoacoustic Imaging with a 2-D CMUT Array. Proceedings. IEEE Ultrasonics Symposium Ma, T., Kothapalli, S. R., Vaithilingam, S., Oralkan, O., Kamaya, A., Wygant, I. O., Zhuang, X., Gambhir, S. S., Jeffrey, R. B., Khuri-Yakub, B. T. 2010; 2010: 375-377

  Abstract

  In this work, we demonstrate 3-D photoacoustic imaging of optically absorbing targets embedded as deep as 5 cm inside a highly scattering background medium using a 2-D capacitive micromachined ultrasonic transducer (CMUT) array with a center frequency of 5.5 MHz. 3-D volumetric images and 2-D maximum intensity projection images are presented to show the objects imaged at different depths. Due to the close proximity of the CMUT to the integrated frontend circuits, the CMUT array imaging system has a low noise floor. This makes the CMUT a promising technology for deep tissue photoacoustic imaging.

  View details for PubMedID 22977296

• Classical Hodgkin Lymphoma in First Complete Remission: Is There a Role for F-18 FDG PET/CT Surveillance? 23rd Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Lagaru, A., Maeda, L. S., Lin, F. I., Hoppe, R. T., Rosenberg, S. A., Gambhir, S. S., Advani, R. H. SPRINGER. 2010: S212–S213

  View details for Web of Science ID 000283023800065

• Combined F-18 Fluoride and F-18 FDG PET/CT Scan for Evaluation of Malignancy: Beyond the Pilot Phase Study 23rd Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Iagaru, A., Mittra, E., Dick, D. W., Gambhir, S. S. SPRINGER. 2010: S200–S200

  View details for Web of Science ID 000283023800010

• Tumor Measurements by F-18-FDG PET: How Accurate are they? 23rd Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Mittra, E., Iagaru, A., Gambhir, S. S. SPRINGER. 2010: S330–S331

  View details for Web of Science ID 000283023800598

• A Comparison Between Time Domain and Spectral Imaging Systems for Imaging Quantum Dots in Small Living Animals MOLECULAR IMAGING AND BIOLOGY de la Zerda, A., Bodapati, S., Teed, R., Schipper, M. L., Keren, S., Smith, B. R., Ng, J. S., Gambhir, S. S. 2010; 12 (5): 500-508

  Abstract

  We quantified the performance of time-domain imaging (TDI) and spectral imaging (SI) for fluorescence imaging of quantum dots (QDs) in three distinct imaging instruments: eXplore Optix (TDI, Advanced Research Technologies Inc.), Maestro (SI, CRi Inc.), and IVIS-Spectrum (SI, Caliper Life Sciences Inc.).The instruments were compared for their sensitivity in phantoms and living mice, multiplexing capabilities (ability to resolve the signal of one QD type in the presence of another), and the dependence of contrast and spatial resolution as a function of depth.In phantoms, eXplore Optix had an order of magnitude better sensitivity compared to the SI systems, detecting QD concentrations of ~40 pM in vitro. Maestro was the best instrument for multiplexing QDs. Reduction of contrast and resolution as a function of depth was smallest with eXplore Optix for depth of 2-6 mm, while other depths gave comparable results in all systems. Sensitivity experiments in living mice showed that the eXplore Optix and Maestro systems outperformed the IVIS-Spectrum.TDI was found to be an order of magnitude more sensitive than SI at the expense of speed and very limited multiplexing capabilities. For deep tissue QD imaging, TDI is most applicable for depths between 2 and 6 mm, as its contrast and resolution degrade the least at these depths.

  View details for DOI 10.1007/s11307-009-0290-4

  View details for Web of Science ID 000282273200006

  View details for PubMedID 20012220

  View details for PubMedCentralID PMC3089652

• [F-18]FPPRGD2 PET/CT Imaging of Integrin Expression in Healthy Volunteers 23rd Annual Congress of the European-Association-of-Nuclear-Medicine (EANM) Mittra, E., Iagaru, A., Goris, M. L., Chin, F., Chen, X., Gambhir, S. S. SPRINGER. 2010: S287–S287

  View details for Web of Science ID 000283023800400

• Noninvasive molecular imaging of c-Myc activation in living mice PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Fan-Minogue, H., Cao, Z., Paulmurugan, R., Chan, C. T., Massoud, T. F., Felsher, D. W., Gambhir, S. S. 2010; 107 (36): 15892-15897

  Abstract

  The cytoplasmic Myc protein (c-Myc) regulates various human genes and is dysregulated in many human cancers. Phosphorylation mediates the protein activation of c-Myc and is essential for the function of this transcription factor in normal cell behavior and tumor growth. To date, however, the targeting of Myc as a therapeutic approach for cancer treatment has been achieved primarily at the nonprotein level. We have developed a molecular imaging sensor for noninvasive imaging of c-Myc activity in living subjects using a split Firefly luciferase (FL) complementation strategy to detect and quantify the phosphorylation-mediated interaction between glycogen synthase kinase 3beta (GSK3beta) and c-Myc. This sensor system consists of two fusion proteins, GSK 35-433-CFL and NFL-c-Myc, in which specific fragments of GSK3beta and c-Myc are fused with C-terminal and N-terminal fragments of the split FL, respectively. The sensor detects phosphorylation-specific GSK3beta-c-Myc interaction, the imaging signal of which correlates with the steady-state and temporal regulation of c-Myc phosphorylation in cell culture. The sensor also detects inhibition of c-Myc activity via differential pathways, allowing noninvasive monitoring of c-Myc-targeted drug efficacy in intact cells and living mice. Notably, this drug inhibition is detected before changes in tumor size are apparent in mouse xenograft and liver tumor models. This reporter system not only provides an innovative way to investigate the role of functional c-Myc in normal and cancer-related biological processes, but also facilitates c-Myc-targeted drug development by providing a rapid quantitative approach to assessing cancer response to therapy in living subjects.

  View details for DOI 10.1073/pnas.1007443107

  View details for PubMedID 20713710

• Biodistribution of Neural Stem Cells After Intravascular Therapy for Hypoxic-Ischemia STROKE Pendharkar, A. V., Chua, J. Y., Andres, R. H., Wang, N., Gaeta, X., Wang, H., De, A., Choi, R., Chen, S., Rutt, B. K., Gambhir, S. S., Guzman, R. 2010; 41 (9): 2064-2070

  Abstract

  Intravascular transplantation of neural stem cells represents a minimally invasive therapeutic approach for the treatment of central nervous system diseases. The cellular biodistribution after intravascular injection needs to be analyzed to determine the ideal delivery modality. We studied the biodistribution and efficiency of targeted central nervous system delivery comparing intravenous and intra-arterial (IA) administration of neural stem cells after brain ischemia.Mouse neural stem cells were transduced with a firefly luciferase reporter gene for bioluminescence imaging (BLI). Hypoxic-ischemia was induced in adult mice and reporter neural stem cells were transplanted IA or intravenous at 24 hours after brain ischemia. In vivo BLI was used to track transplanted cells up to 2 weeks after transplantation and ex vivo BLI was used to determine single organ biodistribution.Immediately after transplantation, BLI signal from the brain was 12 times higher in IA versus intravenous injected animals (P<0.0001). After IA injection, 69% of the total luciferase activity arose from the brain early after transplantation and 93% at 1 week. After intravenous injection, 94% of the BLI signal was detected in the lungs (P=0.004) followed by an overall 94% signal loss at 1 week, indicating lack of cell survival outside the brain. Ex vivo single organ analysis showed a significantly higher BLI signal in the brain than in the lungs, liver, and kidneys at 1 week (P<0.0001) and 2 weeks in IA (P=0.007).IA transplantation results in superior delivery and sustained presence of neural stem cells in the ischemic brain in comparison to intravenous infusion.

  View details for DOI 10.1161/STROKEAHA.109.575993

  View details for PubMedID 20616329

• DYNAMIC CONTRAST-ENHANCED MRI REVEALS CORE SIGNALLING PATHWAYS IN BREAST CANCER 1st British Breast Cancer Research Conference Mehta, S., HUGHES, N. P., Buffa, F. M., Adams, R. F., Gambhir, S. S., Harris, A. L. PERGAMON-ELSEVIER SCIENCE LTD. 2010: 13–13

  View details for DOI 10.1016/j.ejcsup.2010.06.035

  View details for Web of Science ID 000288598400035

• Design, Synthesis, and Imaging of an Activatable Photoacoustic Probe JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Levi, J., Kothapalli, S. R., Ma, T., Hartman, K., Khuri-Yakub, B. T., Gambhir, S. S. 2010; 132 (32): 11264-11269

  Abstract

  Photoacoustic tomography is a rapidly growing imaging modality that can provide images of high spatial resolution and high contrast at depths up to 5 cm. We report here the design, synthesis, and evaluation of an activatable probe that shows great promise for enabling detection of the cleaved probe in the presence of high levels of nonactivated, uncleaved probe, a difficult task to attain in absorbance-based modality. Before the cleavage by its target, proteolytic enzyme MMP-2, the probe, an activatable cell-penetrating peptide, Ceeee[Ahx]PLGLAGrrrrrK, labeled with two chromophores, BHQ3 and Alexa750, shows photoacoustic signals of similar intensity at the two wavelengths corresponding to the absorption maxima of the chromophores, 675 and 750 nm. Subtraction of the images taken at these two wavelengths makes the probe effectively photoacoustically silent, as the signals at these two wavelengths essentially cancel out. After the cleavage, the dye associated with the cell-penetrating part of the probe, BHQ3, accumulates in the cells, while the other dye diffuses away, resulting in photoacoustic signal seen at only one of the wavelengths, 675 nm. Subtraction of the photoacoustic images at two wavelengths reveals the location of the cleaved (activated) probe. In the search for the chromophores that are best suited for photoacoustic imaging, we have investigated the photoacoustic signals of five chromophores absorbing in the near-infrared region. We have found that the photoacoustic signal did not correlate with the absorbance and fluorescence of the molecules, as the highest photoacoustic signal arose from the least absorbing quenchers, BHQ3 and QXL 680.

  View details for DOI 10.1021/0104000a

  View details for Web of Science ID 000280861300058

  View details for PubMedID 20698693

  View details for PubMedCentralID PMC2922742

• [F-18]FTC-146: A novel and highly selective PET ligand for visualizing sigma-1 receptors in living subjects 8th International Symposium on Functional Neuroreceptor Mapping of the Living Brain James, M. L., Shen, B., Zavaleta, C., Berganos, R. A., Mesangeau, C., Shaikh, J., Gambhir, S. S., Matsumoto, R. R., McCurdy, C. R., Chin, F. T. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2010: S123–S124

  View details for DOI 10.1016/j.neuroimage.2010.04.101

  View details for Web of Science ID 000280181900103

• Antiangiogenic Cancer Therapy: Monitoring with Molecular US and a Clinically Translatable Contrast Agent (BR55) RADIOLOGY Pysz, M. A., Foygel, K., Rosenberg, J., Gambhir, S. S., Schneider, M., Willmann, J. K. 2010; 256 (2): 519-527

  Abstract

  To develop and test human kinase insert domain receptor (KDR)-targeted microbubbles (MBs) (MB(KDR)) for imaging KDR at the molecular level and for monitoring antiangiogenic therapy in a human colon cancer xenograft tumor model in mice.Animal studies were approved by the Institutional Administrative Panel on Laboratory Animal Care. A heterodimeric peptide that binds to human KDR with low nanomolar affinity (K(D) = 0.5 nmol/L) was coupled onto the surface of perfluorobutane-containing lipid-shelled MBs (MB(KDR)). Binding specificity of MB(KDR) to human KDR and cross-reactivity with murine vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) were tested in cell culture under flow shear stress conditions (at 100 sec(-1)). In vivo binding specificity of MB(KDR) to VEGFR2 was tested in human LS174T colon cancer xenografts in mice with a 40-MHz ultrasonographic (US) transducer. Targeted contrast material-enhanced US imaging signal by using MB(KDR) was longitudinally measured during 6 days in tumors with (n = 6) and without (n = 6) antiangiogenic treatment (anti-VEGF antibody). Ex vivo VEGFR2 staining and microvessel density analysis were performed. Significant differences were evaluated (t, Mann-Whitney, or Wilcoxon test).Cell culture experiments showed four times greater binding specificity of MB(KDR) to human KDR and cross-reactivity to murine VEGFR2 (P < or = .01). In vivo imaging signal was more than three times higher (P = .01) with MB(KDR) compared with control MBs and decreased significantly (approximately fourfold lower, P = .03) following in vivo receptor blocking with anti-VEGFR2 antibody. One day after initiation of antiangiogenic therapy, imaging signal was significantly decreased (approximately 46% lower, P = .02) in treated versus untreated tumors; it remained significantly lower (range, 46%-84% decreased; P = .038) during the following 5 days. Microvessel density was significantly reduced (P = .04) in treated (mean, 7.3 microvessels per square millimeter +/- 4.7 [standard deviation]) versus untreated tumors (mean, 22.0 microvessels per square millimeter +/- 9.4); VEGFR2 expression was significantly decreased (>50% lower, P = .03) in treated tumors.Human MB(KDR) allow in vivo imaging and longitudinal monitoring of VEGFR2 expression in human colon cancer xenografts.

  View details for DOI 10.1148/radiol.10091858

  View details for Web of Science ID 000280272100023

  View details for PubMedID 20515975

  View details for PubMedCentralID PMC2909432

• A molecularly engineered split reporter for imaging protein-protein interactions with positron emission tomography NATURE MEDICINE Massoud, T. F., Paulmurugan, R., Gambhir, S. S. 2010; 16 (8): 921-U123

  Abstract

  Improved techniques to noninvasively image protein-protein interactions (PPIs) are essential. We molecularly engineered a positron emission tomography (PET)-based split reporter (herpes simplex virus type 1 thymidine kinase), cleaved between Thr265 and Ala266, and used this in a protein-fragment complementation assay (PCA) to quantify PPIs in mammalian cells and to microPET image them in living mice. An introduced point mutation (V119C) markedly enhanced thymidine kinase complementation in PCAs, on the basis of rapamycin modulation of FKBP12-rapamycin-binding domain (FRB) and FKBP12 (FK506 binding protein), the interaction of hypoxia-inducible factor-1alpha with the von Hippel-Lindau tumor suppressor, and in an estrogen receptor intramolecular protein folding assay. Applications of this unique split thymidine kinase are potentially far reaching, including, for example, considerably more accurate monitoring of immune and stem cell therapies, allowing for fully quantitative and tomographic PET localization of PPIs in preclinical small- and large-animal models of disease.

  View details for DOI 10.1038/nm.2185

  View details for Web of Science ID 000280649200033

  View details for PubMedID 20639890

  View details for PubMedCentralID PMC2917476

• Facile Synthesis, Silanization, and Biodistribution of Biocompatible Quantum Dots SMALL Ma, N., Marshall, A. F., Gambhir, S. S., Rao, J. 2010; 6 (14): 1520-1528

  Abstract

  A facile strategy for the synthesis of silica-coated quantum dots (QDs) for in vivo imaging is reported. All the QD synthesis and silanization steps are conducted in water and methanol under mild conditions without involving any organometallic precursors or high-temperature, oxygen-free environments. The as-prepared silica-coated QDs possess high quantum yields and are extremely stable in mouse serum. In addition, the silanization method developed here produces nanoparticles with small sizes that are difficult to achieve via conventional silanization methods. The silica coating helps to prevent the exposure of the QD surface to the biological milieu and therefore increases the biocompatibility of QDs for in vivo applications. Interestingly, the silica-coated QDs exhibit a different biodistribution pattern from that of commercially available Invitrogen QD605 (carboxylate) with a similar size and emission wavelength. The Invitrogen QD605 exhibits predominant liver (57.2% injected dose (ID) g(-1)) and spleen (46.1% ID g(-1)) uptakes 30 min after intravenous injection, whereas the silica-coated QDs exhibit much lower liver (16.2% ID g(-1)) and spleen (3.67% ID g(-1)) uptakes but higher kidney uptake (8.82% ID g(-1)), blood retention (15.0% ID g(-1)), and partial renal clearance. Overall, this straightforward synthetic strategy paves the way for routine and customized synthesis of silica-coated QDs for biological use.

  View details for DOI 10.1002/smll.200902409

  View details for Web of Science ID 000280633900011

  View details for PubMedID 20564726

• Indirect imaging of cardiac-specific transgene expression using a bidirectional two-step transcriptional amplification strategy GENE THERAPY Chen, I. Y., Gheysens, O., Ray, S., Wang, Q., Padmanabhan, P., Paulmurugan, R., Loening, A. M., Rodriguez-Porcel, M., Willmann, J. K., Sheikh, A. Y., Nielsen, C. H., Hoyt, G., Contag, C. H., Robbins, R. C., Biswal, S., Wu, J. C., Gambhir, S. S. 2010; 17 (7): 827-838

  Abstract

  Transcriptional targeting for cardiac gene therapy is limited by the relatively weak activity of most cardiac-specific promoters. We have developed a bidirectional plasmid vector, which uses a two-step transcriptional amplification (TSTA) strategy to enhance the expression of two optical reporter genes, firefly luciferase (fluc) and Renilla luciferase (hrluc), driven by the cardiac troponin T (cTnT) promoter. The vector was characterized in vitro and in living mice using luminometry and bioluminescence imaging to assess its ability to mediate strong, correlated reporter gene expression in a cardiac cell line and the myocardium, while minimizing expression in non-cardiac cell lines and the liver. In vitro, the TSTA system significantly enhanced cTnT-mediated reporter gene expression with moderate preservation of cardiac specificity. After intramyocardial and hydrodynamic tail vein delivery of an hrluc-enhanced variant of the vector, long-term fluc expression was observed in the heart, but not in the liver. In both the cardiac cell line and the myocardium, fluc expression correlated well with hrluc expression. These results show the vector's ability to effectively amplify and couple transgene expression in a cardiac-specific manner. Further replacement of either reporter gene with a therapeutic gene should allow non-invasive imaging of targeted gene therapy in living subjects.

  View details for DOI 10.1038/gt.2010.30

  View details for Web of Science ID 000279614600002

  View details for PubMedID 20237511

  View details for PubMedCentralID PMC2900530

• Molecular imaging: current status and emerging strategies CLINICAL RADIOLOGY Pysz, M. A., Gambhir, S. S., Willmann, J. K. 2010; 65 (7): 500-516

  Abstract

  In vivo molecular imaging has a great potential to impact medicine by detecting diseases in early stages (screening), identifying extent of disease, selecting disease- and patient-specific treatment (personalized medicine), applying a directed or targeted therapy, and measuring molecular-specific effects of treatment. Current clinical molecular imaging approaches primarily use positron-emission tomography (PET) or single photon-emission computed tomography (SPECT)-based techniques. In ongoing preclinical research, novel molecular targets of different diseases are identified and, sophisticated and multifunctional contrast agents for imaging these molecular targets are developed along with new technologies and instrumentation for multi-modality molecular imaging. Contrast-enhanced molecular ultrasound (US) with molecularly-targeted contrast microbubbles is explored as a clinically translatable molecular imaging strategy for screening, diagnosing, and monitoring diseases at the molecular level. Optical imaging with fluorescent molecular probes and US imaging with molecularly-targeted microbubbles are attractive strategies as they provide real-time imaging, are relatively inexpensive, produce images with high spatial resolution, and do not involve exposure to ionizing irradiation. Raman spectroscopy/microscopy has emerged as a molecular optical imaging strategy for ultrasensitive detection of multiple biomolecules/biochemicals with both in vivo and ex vivo versatility. Photoacoustic imaging is a hybrid of optical and US techniques involving optically-excitable molecularly-targeted contrast agents and quantitative detection of resulting oscillatory contrast agent movement with US. Current preclinical findings and advances in instrumentation, such as endoscopes and microcatheters, suggest that these molecular imaging methods have numerous potential clinical applications and will be translated into clinical use in the near future.

  View details for DOI 10.1016/j.crad.2010.03.011

  View details for Web of Science ID 000280379900002

  View details for PubMedID 20541650

  View details for PubMedCentralID PMC3150531

• Reply to: The diagnostic accuracy of F-18-FDG PET in cutaneous malignant melanoma EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Delgado-Bolton, R. C., Jimenez-Requena, F., Fernandez-Perez, C., Gambhir, S. S., Schwimmer, J., Perez-Vazquez, J. M., Carreras-Delgado, J. L. 2010; 37 (7): 1436-1437

  View details for DOI 10.1007/s00259-010-1476-3

  View details for Web of Science ID 000278833100023

• Implantable semiconductor biosensor for continuous in vivo sensing of far-red fluorescent molecules OPTICS EXPRESS O'Sullivan, T., Munro, E. A., Parashurama, N., Conca, C., Gambhir, S. S., Harris, J. S., Levi, O. 2010; 18 (12): 12513-12525

  Abstract

  We have fabricated miniature implantable fluorescence sensors for continuous fluorescence sensing applications in living subjects. These monolithically integrated GaAs-based sensors incorporate a 675 nm vertical-cavity surface-emitting laser (VCSEL), a GaAs PIN photodiode, and a fluorescence emission filter. We demonstrate high detection sensitivity for Cy5.5 far-red dye (50 nanoMolar) in living tissue, limited by the intrinsic background autofluorescence. These low cost, sensitive and scalable sensors are promising for long-term continuous monitoring of molecular dynamics for biomedical studies in freely moving animals.

  View details for Web of Science ID 000278527700052

  View details for PubMedID 20588377

• Cu-64-Labeled Affibody Molecules for Imaging of HER2 Expressing Tumors MOLECULAR IMAGING AND BIOLOGY Cheng, Z., De Jesus, O. P., Kramer, D. J., De, A., Webster, J. M., Gheysens, O., Levi, J., Namavari, M., Wang, S., Park, J. M., Zhang, R., Liu, H., Lee, B., Syud, F. A., Gambhir, S. S. 2010; 12 (3): 316-324

  Abstract

  The development of molecular probes based on novel engineered protein constructs is under active investigation due to the great potential of this generalizable strategy for imaging a variety of tumor targets.In this report, human epidermal growth factor receptor type 2 (HER2)-binding Affibody molecules were radiolabeled with (64)Cu and their imaging ability was further evaluated in tumor mice models to understand the promise and limitations of such probes. The anti-HER2 Affibody molecules in monomeric (Z(HER2:477)) and dimeric [(Z(HER2:477))(2)] forms were site specifically modified with the maleimide-functionalized chelator, 1,4,7,10-tetraazacyclododecane-1,4,7-tris(acetic acid)-10-acetate mono (N-ethylmaleimide amide) (Mal-DOTA). The resulting DOTA-Affibody conjugates were radiolabeled with (64)Cu and evaluated in nude mice bearing subcutaneous SKOV3 tumors. Biodistribution experiments showed that tumor uptake values of (64)Cu-DOTA-Z(HER2:477) and (64)Cu-DOTA-(Z(HER2:477))(2) were 6.12 +/- 1.44% and 1.46 +/- 0.50% ID/g, respectively, in nude mice (n = 3 each) at 4 h postinjection. Moreover, (64)Cu-labeled monomer exhibited significantly higher tumor/blood ratio than that of radiolabeled dimeric counterpart at all time points examined in this study. MicroPET imaging of (64)Cu-DOTA-Z(HER2:477) in SKOV3 tumor mice clearly showed good and specific tumor localization. This study demonstrates that (64)Cu-labeled Z(HER2:477) is a promising targeted molecular probe for imaging HER2 receptor expression in living mice. Further work is needed to improve the excretion properties, hence dosimetry and imaging efficacy, of the radiometal-based probe.

  View details for DOI 10.1007/s11307-009-0256-6

  View details for Web of Science ID 000277375300010

  View details for PubMedID 19779897

• Ultrahigh Sensitivity Carbon Nanotube Agents for Photoacoustic Molecular Imaging in Living Mice NANO LETTERS de la Zerda, A., Liu, Z., Bodapati, S., Teed, R., Vaithilingam, S., Khuri-Yakub, B. T., Chen, X., Dai, H., Gambhir, S. S. 2010; 10 (6): 2168-2172

  Abstract

  Photoacoustic imaging is an emerging modality that overcomes to a great extent the resolution and depth limitations of optical imaging while maintaining relatively high-contrast. However, since many diseases will not manifest an endogenous photoacoustic contrast, it is essential to develop exogenous photoacoustic contrast agents that can target diseased tissue(s). Here we present a novel photoacoustic contrast agent, Indocyanine Green dye-enhanced single walled carbon nanotube (SWNT-ICG). We conjugated this contrast agent with cyclic Arg-Gly-Asp (RGD) peptides to molecularly target the alpha(v)beta(3) integrins, which are associated with tumor angiogenesis. Intravenous administration of this tumor-targeted contrast agent to tumor-bearing mice showed significantly higher photoacoustic signal in the tumor than in mice injected with the untargeted contrast agent. The new contrast agent gave a markedly 300 times higher photoacoustic contrast in living tissues than previously reported SWNTs, leading to subnanomolar sensitivities. Finally, we show that the new contrast agent can detect approximately 20 times fewer cancer cells than previously reported SWNTs.

  View details for DOI 10.1021/nl100890d

  View details for Web of Science ID 000278449200033

  View details for PubMedID 20499887

  View details for PubMedCentralID PMC2893026

• Antioxidants Improve Early Survival of Cardiomyoblasts After Transplantation to the Myocardium MOLECULAR IMAGING AND BIOLOGY Rodriguez-Porcel, M., Gheysens, O., Paulmurugan, R., Chen, I. Y., Peterson, K. M., Willmann, J. K., Wu, J. C., Zhu, X., Lerman, L. O., Gambhir, S. S. 2010; 12 (3): 325-334

  Abstract

  We tested the hypothesis that modulation of the microenvironment (using antioxidants) will increase stem cell survival in hypoxia and after transplantation to the myocardium.Rat cardiomyoblasts were stably transfected with a reporter gene (firefly luciferase) for bioluminescence imaging (BLI). First, we examined the role of oxidative stress in cells under hypoxic conditions. Subsequently, stem cells were transplanted to the myocardium of rats using high-resolution ultrasound, and their survival was monitored daily using BLI.Under hypoxia, oxidative stress was increased together with decreased cell survival compared to control cells, both of which were preserved by antioxidants. In living subjects, oxidative stress blockade increased early cell survival after transplantation to the myocardium, compared to untreated cells/animals.Modulation of the local microenvironment (with antioxidants) improves stem cell survival. Increased understanding of the interaction between stem cells and their microenvironment will be critical to advance the field of regenerative medicine.

  View details for DOI 10.1007/s11307-009-0274-4

  View details for Web of Science ID 000277375300011

  View details for PubMedID 20013064

  View details for PubMedCentralID PMC2865580

• Radiation-Luminescence-Excited Quantum Dots for in vivo Multiplexed Optical Imaging SMALL Liu, H., Zhang, X., Xing, B., Han, P., Gambhir, S. S., Cheng, Z. 2010; 6 (10): 1087-1091

  View details for DOI 10.1002/smll.200902408

  View details for Web of Science ID 000278629300004

  View details for PubMedID 20473988

• Assessing delivery and quantifying efficacy of small interfering ribonucleic acid therapeutics in the skin using a dual-axis confocal microscope JOURNAL OF BIOMEDICAL OPTICS Ra, H., Gonzalez-Gonzalez, E., Smith, B. R., Gambhir, S. S., Kino, G. S., Solgaard, O., Kaspar, R. L., Contag, C. H. 2010; 15 (3)

  Abstract

  Transgenic reporter mice and advances in imaging instrumentation are enabling real-time visualization of cellular mechanisms in living subjects and accelerating the development of novel therapies. Innovative confocal microscope designs are improving their utility for microscopic imaging of fluorescent reporters in living animals. We develop dual-axis confocal (DAC) microscopes for such in vivo studies and create mouse models where fluorescent proteins are expressed in the skin for the purpose of advancing skin therapeutics and transdermal delivery tools. Three-dimensional image volumes, through the different skin compartments of the epidermis and dermis, can be acquired in several seconds with the DAC microscope in living mice, and are comparable to histologic analyses of reporter protein expression patterns in skin sections. Intravital imaging with the DAC microscope further enables visualization of green fluorescent protein (GFP) reporter gene expression in the skin over time, and quantification of transdermal delivery of small interfering RNA (siRNA) and therapeutic efficacy. Visualization of transdermal delivery of nucleic acids will play an important role in the development of innovative strategies for treating skin pathologies.

  View details for DOI 10.1117/1.3432627

  View details for Web of Science ID 000280642900042

  View details for PubMedID 20615029

  View details for PubMedCentralID PMC2904026

• First in man studies of [18F]FPPRGD2: A novel PET radiopharmaceutical for imaging alpha v beta 3 integrin levels Mittra, E., Goris, M., Iagaru, A., Kardan, A., Liu, S., Shen, B., Chin, F., Chen, X., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2010

  View details for Web of Science ID 000447387101008

• Noninvasive molecular imaging of radioactive tracers using optical imaging techniques Liu, H., Ren, G., Miao, Z., Zhang, X., Tang, X., Han, P., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2010

  View details for Web of Science ID 000447387100002

• In vivo multiplexed optical imaging with radiation luminescence excited quantum dots Liu, H., Zhang, X., Xing, B., Han, P., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2010

  View details for Web of Science ID 000447387100003

• Embryonic Stem Cell-Derived Endothelial Cells Engraft Into the Ischemic Hindlimb and Restore Perfusion ARTERIOSCLEROSIS THROMBOSIS AND VASCULAR BIOLOGY Huang, N. F., Niiyama, H., Peter, C., De, A., Natkunam, Y., Fleissner, F., Li, Z., Rollins, M. D., Wu, J. C., Gambhir, S. S., Cooke, J. P. 2010; 30 (5): 984-U224

  Abstract

  We examined the effect of delivery modality on the survival, localization, and functional effects of exogenously administered embryonic stem cells (ESCs) or endothelial cells derived from them (ESC-ECs) in the ischemic hindlimb.Murine ESCs or ESC-ECs were stably transduced with a construct for bioluminescence imaging (BLI) and fluorescent detection. In a syngeneic murine model of limb ischemia, ESCs or ESC-ECs were delivered by intramuscular (IM), intrafemoral artery (IA), or intrafemoral vein injections (n=5 in each group). For 2 weeks, cell survival and localization were tracked by BLI and confirmed by immunohistochemistry, and functional improvement was assessed by laser Doppler perfusion. BLI showed that ESCs localized to the ischemic limb after IM or IA, but not after intrafemoral vein administration. Regardless of the route of administration, ESCs were detected outside the hindlimb circulation in the spleen or lungs. ESCs did not improve limb perfusion and generated teratomas. In contrast, ESC-ECs delivered by all 3 modalities localized to the ischemic limb, as assessed by BLI. Most surprisingly, ESC-EC injected intrafemoral vein eventually localized to the ischemic limb after initially lodging in the pulmonary circulation. Immunohistochemical studies confirmed the engraftment of ESC-ECs into the limb vasculature after 2 weeks. Notably, ESC-ECs were not detected in the spleen or lungs after 2 weeks, regardless of route of administration. Furthermore, ESC-ECs significantly improved limb perfusion and neovascularization compared with the parental ESCs or the vehicle control group.In contrast to parental ESCs, ESC-ECs preferentially localized in the ischemic hindlimb by IA, IM, and intrafemoral vein delivery. ESC-ECs engrafted into the ischemic microvasculature, enhanced neovascularization, and improved limb perfusion.

  View details for DOI 10.1161/ATVBAHA.110.202796

  View details for PubMedID 20167654

• Cetuximab-Based Immunotherapy and Radioimmunotherapy of Head and Neck Squamous Cell Carcinoma CLINICAL CANCER RESEARCH Niu, G., Sun, X., Cao, Q., Courter, D., Koong, A., Le, Q., Gambhir, S. S., Chen, X. 2010; 16 (7): 2095-2105

  Abstract

  To show the relationship between antibody delivery and therapeutic efficacy in head and neck cancers, in this study we evaluated the pharmacokinetics and pharmacodynamics of epidermal growth factor receptor (EGFR)-targeted immunotherapy and radioimmunotherapy by quantitative positron emission tomography (PET) imaging.EGFR expression on UM-SCC-22B and SCC1 human head and neck squamous cell cancer (HNSCC) cells were determined by flow cytometry and immunostaining. Tumor delivery and distribution of cetuximab in tumor-bearing nude mice were evaluated with small animal PET using (64)Cu-DOTA-cetuximab. The in vitro toxicity of cetuximab to HNSCC cells was evaluated by MTT assay. The tumor-bearing mice were then treated with four doses of cetuximab at 10 mg/kg per dose, and tumor growth was evaluated by caliper measurement. FDG PET was done after the third dose of antibody administration to evaluate tumor response. Apoptosis and tumor cell proliferation after cetuximab treatment were analyzed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling and Ki-67 staining. Radioimmunotherapy was done with (90)Y-DOTA-cetuximab.EGFR expression on UM-SCC-22B cells is lower than that on SCC1 cells. However, the UM-SCC-22B tumors showed much higher (64)Cu-DOTA-cetuximab accumulation than the SCC1 tumors. Cetuximab-induced apoptosis in SCC1 tumors and tumor growth was significantly inhibited, whereas an agonistic effect of cetuximab on UM-SCC-22B tumor growth was observed. After cetuximab treatment, the SCC1 tumors showed decreased FDG uptake, and the UM-SCC-22B tumors had increased FDG uptake. UM-SCC-22B tumors are more responsive to (90)Y-DOTA-cetuximab treatment than SCC1 tumors, partially due to the high tumor accumulation of the injected antibody.Cetuximab has an agonistic effect on the growth of UM-SCC-22B tumors, indicating that tumor response to cetuximab treatment is not necessarily related to EGFR expression and antibody delivery efficiency, as determined by PET imaging. Although PET imaging with antibodies as tracers has limited function in patient screening, it can provide guidance for targeted therapy using antibodies as delivery vehicles.

  View details for DOI 10.1158/1078-0432.CCR-09-2495

  View details for Web of Science ID 000278595800013

  View details for PubMedID 20215534

  View details for PubMedCentralID PMC2848903

• Molecular imaging of biological gene delivery vehicles for targeted cancer therapy: beyond viral vectors. Nuclear medicine and molecular imaging Min, J., Nguyen, V. H., Gambhir, S. S. 2010; 44 (1): 15-24

  Abstract

  Cancer persists as one of the most devastating diseases in the world. Problems including metastasis and tumor resistance to chemotherapy and radiotherapy have seriously limited the therapeutic effects of present clinical treatments. To overcome these limitations, cancer gene therapy has been developed over the last two decades for a broad spectrum of applications, from gene replacement and knockdown to vaccination, each with different requirements for gene delivery. So far, a number of genes and delivery vectors have been investigated, and significant progress has been made with several gene therapy modalities in clinical trials. Viral vectors and synthetic liposomes have emerged as the vehicles of choice for many applications. However, both have limitations and risks that restrict gene therapy applications, including the complexity of production, limited packaging capacity, and unfavorable immunological features. While continuing to improve these vectors, it is important to investigate other options, particularly nonviral biological agents such as bacteria, bacteriophages, and bacteria-like particles. Recently, many molecular imaging techniques for safe, repeated, and high-resolution in vivo imaging of gene expression have been employed to assess vector-mediated gene expression in living subjects. In this review, molecular imaging techniques for monitoring biological gene delivery vehicles are described, and the specific use of these methods at different steps is illustrated. Linking molecular imaging to gene therapy will eventually help to develop novel gene delivery vehicles for preclinical study and support the development of future human applications.

  View details for DOI 10.1007/s13139-009-0006-3

  View details for PubMedID 24899933

  View details for PubMedCentralID PMC4042968

• Rosiglitazone Increases Myocardial Glucose Metabolism in Insulin-Resistant Cardiomyopathy JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY Kao, D. P., Witteles, R. M., Quon, A., Wu, J. C., Gambhir, S. S., Fowler, M. B. 2010; 55 (9): 926-927

  View details for DOI 10.1016/j.jacc.2009.08.085

  View details for Web of Science ID 000274865100015

  View details for PubMedID 20185047

• Molecular Optical Imaging with Radioactive Probes PLOS ONE Liu, H., Ren, G., Miao, Z., Zhang, X., Tang, X., Han, P., Gambhir, S. S., Cheng, Z. 2010; 5 (3)

  Abstract

  Optical imaging (OI) techniques such as bioluminescence and fluorescence imaging have been widely used to track diseases in a non-invasive manner within living subjects. These techniques generally require bioluminescent and fluorescent probes. Here we demonstrate the feasibility of using radioactive probes for in vivo molecular OI.By taking the advantages of low energy window of light (1.2-3.1 eV, 400-1000 nm) resulting from radiation, radionuclides that emit charged particles such as beta(+) and beta(-) can be successfully imaged with an OI instrument. In vivo optical images can be obtained for several radioactive probes including 2-deoxy-2-[(18)F]fluoro-D-glucose ([(18)F]FDG), Na(18)F, Na(131)I, (90)YCl(3) and a (90)Y labeled peptide that specifically target tumors.These studies demonstrate generalizability of radioactive OI technique. It provides a new molecular imaging strategy and will likely have significant impact on both small animal and clinical imaging.

  View details for DOI 10.1371/journal.pone.0009470

  View details for Web of Science ID 000274997100016

  View details for PubMedID 20208993

  View details for PubMedCentralID PMC2830426

• A Dual-Labeled Knottin Peptide for PET and Near-Infrared Fluorescence Imaging of Integrin Expression in Living Subjects BIOCONJUGATE CHEMISTRY Kimura, R. H., Miao, Z., Cheng, Z., Gambhir, S. S., Cochran, J. R. 2010; 21 (3): 436-444

  Abstract

  Previously, we used directed evolution to engineer mutants of the Ecballium elaterium trypsin inhibitor (EETI-II) knottin that bind to alpha(v)beta(3) and alpha(v)beta(5) integrin receptors with low nanomolar affinity, and showed that Cy5.5- or (64)Cu-DOTA-labeled knottin peptides could be used to image integrin expression in mouse tumor models using near-infrared fluorescence (NIRF) imaging or positron emission tomography (PET). Here, we report the development of a dual-labeled knottin peptide conjugated to both NIRF and PET imaging agents for multimodality imaging in living subjects. We created an orthogonally protected peptide-based linker for stoichiometric coupling of (64)Cu-DOTA and Cy5.5 onto the knottin N-terminus and confirmed that conjugation did not affect binding to alpha(v)beta(3) and alpha(v)beta(5) integrins. NIRF and PET imaging studies in tumor xenograft models showed that Cy5.5 conjugation significantly increased kidney uptake and retention compared to the knottin peptide labeled with (64)Cu-DOTA alone. In the tumor, the dual-labeled (64)Cu-DOTA/Cy5.5 knottin peptide showed decreased wash-out leading to significantly better retention (p < 0.05) compared to the (64)Cu-DOTA-labeled knottin peptide. Tumor uptake was significantly reduced (p < 0.05) when the dual-labeled knottin peptide was coinjected with an excess of unlabeled competitor and when tested in a tumor model with lower levels of integrin expression. Finally, plots of tumor-to-background tissue ratios for Cy5.5 versus (64)Cu uptake were well-correlated over several time points post injection, demonstrating pharmacokinetic cross validation of imaging labels. This dual-modality NIRF/PET imaging agent is promising for further development in clinical applications where high sensitivity and high resolution are desired, such as detection of tumors located deep within the body and image-guided surgical resection.

  View details for DOI 10.1021/bc9003102

  View details for Web of Science ID 000275711600004

  View details for PubMedCentralID PMC3004996

• Targeted Contrast-Enhanced Ultrasound Imaging of Tumor Angiogenesis with Contrast Microbubbles Conjugated to Integrin-Binding Knottin Peptides JOURNAL OF NUCLEAR MEDICINE Willmann, J. K., Kimura, R. H., Deshpande, N., Lutz, A. M., Cochran, J. R., Gambhir, S. S. 2010; 51 (3): 433-440

  Abstract

  Targeted contrast-enhanced ultrasound imaging is increasingly being recognized as a powerful imaging tool for the detection and quantification of tumor angiogenesis at the molecular level. The purpose of this study was to develop and test a new class of targeting ligands for targeted contrast-enhanced ultrasound imaging of tumor angiogenesis with small, conformationally constrained peptides that can be coupled to the surface of ultrasound contrast agents.Directed evolution was used to engineer a small, disulfide-constrained cystine knot (knottin) peptide that bound to alpha(v)beta(3) integrins with a low nanomolar affinity (Knottin(Integrin)). A targeted contrast-enhanced ultrasound imaging contrast agent was created by attaching Knottin(Integrin) to the shell of perfluorocarbon-filled microbubbles (MB-Knottin(Integrin)). A knottin peptide with a scrambled sequence was used to create control microbubbles (MB-Knottin(Scrambled)). The binding of MB-Knottin(Integrin) and MB-Knottin(Scrambled) to alpha(v)beta(3) integrin-positive cells and control cells was assessed in cell culture binding experiments and compared with that of microbubbles coupled to an anti-alpha(v)beta(3) integrin monoclonal antibody (MB(alphavbeta3)) and microbubbles coupled to the peptidomimetic agent c(RGDfK) (MB(cRGD)). The in vivo imaging signals of contrast-enhanced ultrasound with the different types of microbubbles were quantified in 42 mice bearing human ovarian adenocarcinoma xenograft tumors by use of a high-resolution 40-MHz ultrasound system.MB-Knottin(Integrin) attached significantly more to alpha(v)beta(3) integrin-positive cells (1.76 +/- 0.49 [mean +/- SD] microbubbles per cell) than to control cells (0.07 +/- 0.006). Control MB-Knottin(Scrambled) adhered less to alpha(v)beta(3) integrin-positive cells (0.15 +/- 0.12) than MB-Knottin(Integrin). After blocking of integrins, the attachment of MB-Knottin(Integrin) to alpha(v)beta(3) integrin-positive cells decreased significantly. The in vivo ultrasound imaging signal was significantly higher after the administration of MB-Knottin(Integrin) than after the administration of MB(alphavbeta3) or control MB-Knottin(Scrambled). After in vivo blocking of integrin receptors, the imaging signal after the administration of MB-Knottin(Integrin) decreased significantly (by 64%). The imaging signals after the administration of MB-Knottin(Integrin) were not significantly different in the groups of tumor-bearing mice imaged with MB-Knottin(Integrin) and with MB(cRGD). Ex vivo immunofluorescence confirmed integrin expression on endothelial cells of human ovarian adenocarcinoma xenograft tumors.Integrin-binding knottin peptides can be conjugated to the surface of microbubbles and used for in vivo targeted contrast-enhanced ultrasound imaging of tumor angiogenesis. Our results demonstrate that microbubbles conjugated to small peptide-targeting ligands provide imaging signals higher than those provided by a large antibody molecule.

  View details for DOI 10.2967/jnumed.109.068007

  View details for Web of Science ID 000275133100026

  View details for PubMedID 20150258

• Evaluation of a Cu-64-Labeled Cystine-Knot Peptide Based on Agouti-Related Protein for PET of Tumors Expressing alpha(v)beta(3) Integrin JOURNAL OF NUCLEAR MEDICINE Jiang, L., Kimura, R. H., Miao, Z., Silverman, A. P., Ren, G., Liu, H., Li, P., Gambhir, S. S., Cochran, J. R., Cheng, Z. 2010; 51 (2): 251-258

  Abstract

  Recently, a truncated form of the agouti-related protein (AgRP), a 4-kDa cystine-knot peptide of human origin, was used as a scaffold to engineer mutants that bound to alpha(v)beta(3) integrin with high affinity and specificity. In this study, we evaluated the potential of engineered integrin-binding AgRP peptides for use as cancer imaging agents in living subjects.Engineered AgRP peptides were prepared by solid-phase peptide synthesis and were folded in vitro and purified by reversed-phase high-performance liquid chromatography. Competition assays were used to measure the relative binding affinities of engineered AgRP peptides for integrin receptors expressed on the surface of U87MG glioblastoma cells. The highest-affinity mutant, AgRP clone 7C, was site-specifically conjugated with 1,4,7,10-tetra-azacyclododecane-N,N',N''N'''-tetraacetic acid (DOTA). The resulting bioconjugate, DOTA-AgRP-7C, was radiolabeled with (64)Cu for biodistribution analysis and small-animal PET studies in mice bearing U87MG tumor xenografts. In addition to serum stability, the in vivo metabolic stability of (64)Cu-DOTA-AgRP-7C was assessed after injection and probe recovery from mouse kidney, liver, tumor, and urine.AgRP-7C and DOTA-AgRP-7C bound with high affinity to integrin receptors expressed on U87MG cells (half maximal inhibitory concentration values, 20 +/- 4 and 14 +/- 2 nM, respectively). DOTA-AgRP-7C was labeled with (64)Cu with high radiochemical purity (>99%). In biodistribution and small-animal PET studies, (64)Cu-DOTA-AgRP-7C displayed rapid blood clearance, good tumor uptake and retention (2.70 +/- 0.93 percentage injected dose per gram [%ID/g] and 2.37 +/- 1.04 %ID/g at 2 and 24 h, respectively), and high tumor-to-background tissue ratios. The integrin-binding specificity of (64)Cu-DOTA-AgRP-7C was confirmed in vitro and in vivo by showing that a large molar excess of the unlabeled peptidomimetic c(RGDyK) could block probe binding and tumor uptake. Serum stability and in vivo metabolite assays demonstrated that engineered AgRP peptides are sufficiently stable for in vivo molecular imaging applications.A radiolabeled version of the engineered AgRP peptide 7C showed promise as a PET agent for tumors that express the alpha(v)beta(3) integrin. Collectively, these results validate AgRP-based cystine-knot peptides for use in vivo as molecular imaging agents and provide support for the general use of AgRP as a scaffold to develop targeting peptides, and hence diagnostics, against other tumor receptors.

  View details for DOI 10.2967/jnumed.109.069831

  View details for Web of Science ID 000274152800028

  View details for PubMedID 20124048

• A Dual-Labeled Knottin Peptide for PET and Near-Infrared Fluorescence Imaging of Integrin Expression in Living Subjects. Bioconjugate chemistry Kimura, R. H., Miao, Z., Cheng, Z., Gambhir, S. S., Cochran, J. R. 2010

  Abstract

  Previously, we used directed evolution to engineer mutants of the Ecballium elaterium trypsin inhibitor (EETI-II) knottin that bind to alpha(v)beta(3) and alpha(v)beta(5) integrin receptors with low nanomolar affinity, and showed that Cy5.5- or (64)Cu-DOTA-labeled knottin peptides could be used to image integrin expression in mouse tumor models using near-infrared fluorescence (NIRF) imaging or positron emission tomography (PET). Here, we report the development of a dual-labeled knottin peptide conjugated to both NIRF and PET imaging agents for multimodality imaging in living subjects. We created an orthogonally protected peptide-based linker for stoichiometric coupling of (64)Cu-DOTA and Cy5.5 onto the knottin N-terminus and confirmed that conjugation did not affect binding to alpha(v)beta(3) and alpha(v)beta(5) integrins. NIRF and PET imaging studies in tumor xenograft models showed that Cy5.5 conjugation significantly increased kidney uptake and retention compared to the knottin peptide labeled with (64)Cu-DOTA alone. In the tumor, the dual-labeled (64)Cu-DOTA/Cy5.5 knottin peptide showed decreased wash-out leading to significantly better retention (p < 0.05) compared to the (64)Cu-DOTA-labeled knottin peptide. Tumor uptake was significantly reduced (p < 0.05) when the dual-labeled knottin peptide was coinjected with an excess of unlabeled competitor and when tested in a tumor model with lower levels of integrin expression. Finally, plots of tumor-to-background tissue ratios for Cy5.5 versus (64)Cu uptake were well-correlated over several time points post injection, demonstrating pharmacokinetic cross validation of imaging labels. This dual-modality NIRF/PET imaging agent is promising for further development in clinical applications where high sensitivity and high resolution are desired, such as detection of tumors located deep within the body and image-guided surgical resection.

  View details for DOI 10.1021/bc9003102

  View details for PubMedID 20131753

  View details for PubMedCentralID PMC3004996

• Meta-analysis of the performance of F-18-FDG PET in cutaneous melanoma EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Jimenez-Requena, F., Delgado-Bolton, R. C., Fernandez-Perez, C., Gambhir, S. S., Schwimmer, J., Perez-Vazquez, J. M., Carreras-Delgado, J. L. 2010; 37 (2): 284-300

  Abstract

  The aim of this study was to perform a systematic review of the literature to evaluate the accuracy of FDG-PET in staging and restaging of cutaneous melanoma.Systematic methods were used to identify, select, and evaluate the methodologic quality of the studies as well as to summarize the overall findings of sensitivity and specificity. The search strategy consisted of identifying studies published between 2000 and 2006. Inclusion criteria were studies that evaluated the diagnostic performance of FDG-PET in staging/restaging of cutaneous melanoma. The results were compared and pooled with a meta-analysis published previously that included studies published until 1999. The meta-analysis included 95% confidence intervals (CI) of sensitivity, specificity, likelihood-ratio (LR), and diagnostic-odds-ratio (DOR).The quantitative meta-analysis included 24 studies that were analysed in two groups: eight studies were included only in the regional staging analysis (group I), 13 studies were included only in the detection of distant metastases analysis (group II), and three studies were included in both analyses. Compliance with the methodologic-quality criteria was acceptable. We analysed the results of data presented in patients, lesions, basins, lymph-nodes, areas, and scans. Regarding the performance of FDG-PET in the detection of metastases, the pooled studies presented homogeneity for the negative-LR (0.15; 95% CI, 0.10-0.22) when analyzing lesions. When analyzing scans, there was global homogeneity for specificity (0.86; 95% CI, 0.77-0.92), positive-LR (5.86; 95% CI, 3.64-9.43), and DOR (37.89; 95% CI, 15.80-90.86). The pooled studies presented heterogeneity for the other items analysed. Regarding the detection of regional metastases, when analyzing lymph-nodes there was global homogeneity for specificity (0.99; 95% CI, 0.97-0.99; P = 0.101). The meta-regression evidenced that the variable that most influenced the DOR of the different studies and that can explain the heterogeneity was the year of publication; this may be related to the evolution of PET technology and an improvement of sensitivity/specificity.FDG-PET is not useful in the evaluation of regional metastases, as it does not detect microscopic disease. However, FDG-PET could be useful in the detection of distant metastases, and could suggest its utility in the management of patients with cutaneous melanoma.

  View details for DOI 10.1007/s00259-009-1224-8

  View details for Web of Science ID 000274293900011

  View details for PubMedID 19727717

  View details for PubMedCentralID PMC2886141

• Photoacoustic ocular imaging OPTICS LETTERS de la Zerda, A., Paulus, Y. M., Teed, R., Bodapati, S., Dollberg, Y., Khuri-Yakub, B. T., Blumenkranz, M. S., Moshfeghi, D. M., Gambhir, S. S. 2010; 35 (3): 270-272

  Abstract

  We developed a photoacoustic ocular imaging device and demonstrated its utility in imaging the deeper layers of the eye including the retina, choroid, and optic nerve. Using safe laser intensity, the photoacoustic system was able to visualize the blood distribution of an enucleated pig's eye and an eye of a living rabbit. Ultrasound images, which were simultaneously acquired, were overlaid on the photoacoustic images to visualize the eye's anatomy. Such a system may be used in the future for early detection and improved management of neovascular ocular diseases, including wet age-related macular degeneration and proliferative diabetic retinopathy.

  View details for PubMedID 20125691

• A Novel Method of Monitoring Placenta-Specific Transgene Expression Throughout Pregnancy by Noninvasive Bioluminescence Imaging 43rd Annual Meeting of the Society-for-the-Study-of-Reproduction Fan, X., Ren, P., Dhal, S., Goodman, S. B., Gambhir, S. S., Druzin, M. L., Nayak, N. R. SOC STUDY REPRODUCTION. 2010: 144–145

  View details for Web of Science ID 000284381300406

• Molecular Imaging with Reporter Genes Gambhir, S. S., Yaghoubi, S. Cambridge University Press. 2010
• Molecular Imaging: Principles and Practice Weissleder, R., Ross, B., Rehemtulla, A., Gambhir, S. S. People's Medical Publishing House. 2010
• Encyclopedic Reference on Imaging Springer Medicine eBook Collection Chan, C., Gambhir, S. S. edited by Ntziachristos, V. Springer. 2010
• Gene Therapy and Imaging of Transgene Expression in Living Subjects Molecular Imaging with Reporter Genes Gambhir, S. S., Yaghoubi, S. edited by Gambhir, S. S., Yaghoubi, S. Cambridge University Press. 2010: 227–238
• Cell-Specific Imaging of Reporter Gene Expression Using a Two-Step Transcriptional Amplification Strategy Molecular Imaging with Reporter Genes Figueiredo, M. L., Gambhir, S. S., Carey, M., Wu, L. edited by Gambhir, S. S., Yaghoubi, S. Cambridge University Press. 2010: 127–150
• Reporter Gene Imaging of Cell Signal Transduction Molecular Imaging with Reporter Genes Massoud, T. F., Paulmurugan, R., Chan, C., Fan-Minogue, H., Gambhir, S. S. edited by Yaghoubi, S., Gambhir, S. S. Cambridge University Press. 2010: 195–226
• Clinical Applications of Reporter Gene Technology in Living Subjects Molecular Imaging with Reporter Genes Penuelas, I., Yaghoubi, S., Prosper, F., Gambhir, S. S. edited by Yaghoubi, S., Gambhir, S. S. 2010: 297–314
• Imaging of Reporter Genes and Stem Cells Molecular Imaging with Reporter Genes Rodriguez-Porcel, M. G., Gambhir, S. S. edited by Yaghoubi, S., Gambhir, S. S. Cambridge University Press. 2010: 275–296
• Molecular Imaging of Gene Expression and Cell Therapy Clinical Nuclear Cardiology: State of the Art and Future Directions Wu, J., Gambhir, S. S. edited by Zaret, B., Beller, G. Mosby, Inc.. 2010; 4: 723–737
• A red-shifted Renilla luciferase for transient reporter-gene expression NATURE METHODS Loening, A. M., Dragulescu-Andrasi, A., Gambhir, S. S. 2010; 7 (1): 5-6

  View details for DOI 10.1038/nmeth0110-05

  View details for Web of Science ID 000273128300003

  View details for PubMedID 20038949

• Optical Imaging with Radioactive Probes PLoS One Liu, H., Ren G, Miao Z, Zhang X, Tang X, Han P, Gambhir SS, Cheng Z 2010; 5 (3): E9470
• Meta-analysis of the Performance of [18F] FDG-PET in Cutaneous Melanoma. European Journal of Nuclear Medicine and Molecular Imaging JImenez-Requena F, Delgado-Bolton R, Fernandez-Perez C, Gambhir SS, Schwimmer J, Perez-Vazquez J, Carreras-Delgado J. 2010; 37(2): 284-300
• Combined F-18-FDG and Fluoride Approach in PET/CT Imaging: Is There a Clinical Future? REPLY JOURNAL OF NUCLEAR MEDICINE Iagaru, A., Mittra, E., Goris, M. L., Gambhir, S. S. 2010; 51 (1): 166-167

  View details for DOI 10.2967/jnumed.109.067082

  View details for Web of Science ID 000273263800030

• Functional and Transcriptional Characterization of Human Embryonic Stem Cell-Derived Endothelial Cells for Treatment of Myocardial Infarction PLOS ONE Li, Z., Wilson, K. D., Smith, B., Kraft, D. L., Jia, F., Huang, M., Xie, X., Robbins, R. C., Gambhir, S. S., Weissman, I. L., Wu, J. C. 2009; 4 (12)

  Abstract

  Differentiation of human embryonic stem cells into endothelial cells (hESC-ECs) has the potential to provide an unlimited source of cells for novel transplantation therapies of ischemic diseases by supporting angiogenesis and vasculogenesis. However, the endothelial differentiation efficiency of the conventional embryoid body (EB) method is low while the 2-dimensional method of co-culturing with mouse embryonic fibroblasts (MEFs) require animal product, both of which can limit the future clinical application of hESC-ECs. Moreover, to fully understand the beneficial effects of stem cell therapy, investigators must be able to track the functional biology and physiology of transplanted cells in living subjects over time.In this study, we developed an extracellular matrix (ECM) culture system for increasing endothelial differentiation and free from contaminating animal cells. We investigated the transcriptional changes that occur during endothelial differentiation of hESCs using whole genome microarray, and compared to human umbilical vein endothelial cells (HUVECs). We also showed functional vascular formation by hESC-ECs in a mouse dorsal window model. Moreover, our study is the first so far to transplant hESC-ECs in a myocardial infarction model and monitor cell fate using molecular imaging methods.Taken together, we report a more efficient method for derivation of hESC-ECs that express appropriate patterns of endothelial genes, form functional vessels in vivo, and improve cardiac function. These studies suggest that hESC-ECs may provide a novel therapy for ischemic heart disease in the future.

  View details for DOI 10.1371/journal.pone.0008443

  View details for Web of Science ID 000273180200002

  View details for PubMedID 20046878

  View details for PubMedCentralID PMC2795856

• Efficacy of F-18-FDG PET/CT in the evaluation of patients with recurrent cervical carcinoma EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Mittra, E., El-Maghraby, T., Rodriguez, C. A., Quon, A., McDougall, I. R., Gambhir, S. S., Iagaru, A. 2009; 36 (12): 1952-1959

  Abstract

  Only a limited number of studies have evaluated the efficacy of 18F-FDG PET/CT for recurrent cervical carcinoma, which this study seeks to expand upon.This is a retrospective study of 30 women with cervical carcinoma who had a surveillance PET/CT after initial therapy. Sensitivity, specificity, accuracy, positive predictive value, and negative predictive value were calculated using a 2 × 2 contingency table with pathology results (76%) or clinical follow-up (24%) as the gold standard. The Wilson score method was used to perform 95% confidence interval estimations.The sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of PET/CT for the detection of local recurrence at the primary site were 93, 93, 93, 86, and 96%, respectively. The same values for the detection of distant metastases were 96, 95, 95, 96, and 95%, respectively. Seventy-one percent of the scans performed in symptomatic patients showed true-positive findings. In comparison, 44% of scans performed in asymptomatic patients showed true-positive findings. But, all patients subsequently had a change in their management based on the PET/CT findings such that the effect was notable. The maximum standardized uptake value ranged from 5 to 28 (average: 13 ± 7) in the primary site and 3 to 23 (average: 8 ± 4) in metastases which were significantly different (p = 0.04).This study demonstrates favorable efficacy of 18F-FDG PET/CT for identification of residual/recurrent cervical cancer, as well as for localization of distant metastases.

  View details for DOI 10.1007/s00259-009-1206-x

  View details for Web of Science ID 000271979300004

  View details for PubMedID 19585114

• Development and intra-institutional and inter-institutional validation of a comprehensive new hepatobiliary software: part 1-liver and gallbladder function NUCLEAR MEDICINE COMMUNICATIONS Krishnamurthy, G. T., Krishnamurthy, S., Gambhir, S. S., Rodrigues, C., Rosenberg, J., Schiepers, C., Buxton-Thomas, M. 2009; 30 (12): 934-944

  Abstract

  To develop a software tool for quantification of liver and gallbladder function, and to assess the repeatability and reproducibility of measurements made with it.The software tool developed with the JAVA programming language uses the JAVA2 Standard Edition framework. After manual selection of the regions of interest on a 99mTc hepatic iminodiacetic acid study, the program calculates differential hepatic bile flow, basal duodeno-gastric bile reflux (B-DGBR), hepatic extraction fraction (HEF) of both the lobes with deconvolutional analysis and excretion half-time with nonlinear least squares fit. Gallbladder ejection fraction, ejection period (EP), ejection rate (ER), and postcholecystokinin (CCK) DGBR are calculated after stimulation with CCK-8. To assess intra-observer repeatability and intra-observer reproducibility, measurements from 10 normal participants were analyzed twice by three nuclear medicine technologists at the primary center. To assess inter-site reproducibility, measurements from a superset of 24 normal participants were also assessed once by three observers at the primary center and single observer at three other sites.For the 24 control participants, mean+/-SD of hepatic bile flow into gallbladder was 63.87+/-28.7%, HEF of the right lobe 100+/-0%, left lobe 99.43+2.63%, excretion half-time of the right lobe 21.50+6.98 min, left lobe 28.3+/-11.3 min. Basal DGBR was 1.2+/-1.0%. Gallbladder ejection fraction was 80+/-11%, EP 15.0+/-3.0 min, ER 5.8+/-1.6%/min, and DGBR-CCK 1.3+/-2.3%. Left and right lobe HEF was virtually identical across readers. All measures showed high repeatability except for gallbladder bile flow, basal DGBR, and EP, which exhibited marginal repeatability. Ejection fraction exhibited high reproducibility. There was high concordance among the three primary center observers except for basal DGBR, EP, and ER. Concordance between the primary site and one of the other sites was high, one was fair, and one was poor.New United States Food and Drug Administration-approved personal computer-based Krishnamurthy Hepato-Biliary Software for quantification of the liver and gallbladder function shows promise for consistently repeatable and reproducible results both within and between institutions, and may help to promote universal standardization of data acquisition and analysis in nuclear hepatology.

  View details for DOI 10.1097/MNM.0b013e32832ed34a

  View details for Web of Science ID 000272116100006

  View details for PubMedID 19858769

• Embryonic Stem Cell-Derived Endothelial Cells Engraft Into the Ischemic Hindlimb and Restore Perfusion 82nd National Conference and Exhibitions and Scientific Sessions of the American-Heart-Association Huang, N. F., Niiyama, H., Peter, C., De, A., Natkunam, Y., Fleissner, F., Li, Z., Rollins, M. D., Wu, J. C., Gambhir, S. S., Cooke, J. P. LIPPINCOTT WILLIAMS & WILKINS. 2009: S1152–S1152

  View details for Web of Science ID 000271831504251

• A Novel Molecular Imaging Sensor of Cellular Oxidative Stress 82nd National Conference and Exhibitions and Scientific Sessions of the American-Heart-Association Peterson, K. M., Chen, I. Y., Simari, R. D., Gambhir, S. S., Lerman, A., Rodriguez-Porcel, M. LIPPINCOTT WILLIAMS & WILKINS. 2009: S1025–S1025

  View details for Web of Science ID 000271831503510

• F-18-FDG Uptake in Lung, Breast, and Colon Cancers: Molecular Biology Correlates and Disease Characterization JOURNAL OF NUCLEAR MEDICINE Jadvar, H., Alavi, A., Gambhir, S. S. 2009; 50 (11): 1820-1827

  Abstract

  It is hoped that in the not too distant future, noninvasive imaging-based molecular interrogation and characterization of tumors can improve our fundamental understanding of the dynamic biologic behavior of cancer. For example, the new dimension of diagnostic information that is provided by (18)F-FDG PET has led to improved clinical decision making and management changes in a substantial number of patients with cancer. In this context, the aim of this review is to bring together and summarize the current data on the correlation between the underlying molecular biology and the clinical observations of tumor (18)F-FDG accumulation in 3 major human cancers: lung, breast, and colon.

  View details for DOI 10.2967/jnumed.108.054098

  View details for Web of Science ID 000272554100015

  View details for PubMedID 19837767

  View details for PubMedCentralID PMC2783751

• Three-Dimensional Photoacoustic Imaging Using a Two-Dimensional CMUT Array IEEE TRANSACTIONS ON ULTRASONICS FERROELECTRICS AND FREQUENCY CONTROL Vaithilingam, S., Ma, T., Furukawa, Y., Wygant, I. O., Zhuang, X., de la Zerda, A., Oralkan, O., Kamaya, A., Gambhir, S. S., Jeffrey, R. B., Khuri-Yakub, B. T. 2009; 56 (11): 2411-2419

  Abstract

  In this paper, we describe using a 2-D array of capacitive micromachined ultrasonic transducers (CMUTs) to perform 3-D photoacoustic and acoustic imaging. A tunable optical parametric oscillator laser system that generates nanosecond laser pulses was used to induce the photoacoustic signals. To demonstrate the feasibility of the system, 2 different phantoms were imaged. The first phantom consisted of alternating black and transparent fishing lines of 180 mum and 150 mum diameter, respectively. The second phantom comprised polyethylene tubes, embedded in chicken breast tissue, filled with liquids such as the dye indocyanine green, pig blood, and a mixture of the 2. The tubes were embedded at a depth of 0.8 cm inside the tissue and were at an overall distance of 1.8 cm from the CMUT array. Two-dimensional cross-sectional slices and 3-D volume rendered images of pulse-echo data as well as photoacoustic data are presented. The profile and beamwidths of the fishing line are analyzed and compared with a numerical simulation carried out using the Field II ultrasound simulation software. We investigated using a large aperture (64 x 64 element array) to perform photoacoustic and acoustic imaging by mechanically scanning a smaller CMUT array (16 x 16 elements). Two-dimensional transducer arrays overcome many of the limitations of a mechanically scanned system and enable volumetric imaging. Advantages of CMUT technology for photoacoustic imaging include the ease of integration with electronics, ability to fabricate large, fully populated 2-D arrays with arbitrary geometries, wide-bandwidth arrays and high-frequency arrays. A CMUT based photoacoustic system is proposed as a viable alternative to a piezoelectric transducer based photoacoustic systems.

  View details for DOI 10.1109/TUFFC.2009.1329

  View details for Web of Science ID 000271478600010

  View details for PubMedID 19942528

• PET of Malignant Melanoma Using F-18-Labeled Metallopeptides JOURNAL OF NUCLEAR MEDICINE Ren, G., Liu, Z., Miao, Z., Liu, H., Subbarayan, M., Chin, F. T., Zhang, L., Gambhir, S. S., Cheng, Z. 2009; 50 (11): 1865-1872

  Abstract

  Melanocortin type 1 receptor (MC1R), also known as alpha-melanocyte-stimulating hormone (alpha-MSH) receptor, is an attractive molecular target for melanoma imaging and therapy. An (18)F-labeled linear alpha-MSH peptide ((18)F-FB-Ac-Nle-Asp-His-d-Phe-Arg-Trp-Gly-Lys-NH(2) [NAPamide]) shows promising melanoma imaging properties but with only moderate tumor uptake and retention. A transition metal rhenium-cyclized alpha-MSH peptide, ReO[Cys(3,4,10),d-Phe(7),Arg(11)]alpha-MSH(3-13) (ReCCMSH(Arg(11))), has shown high in vitro binding affinity to MC1R and excellent in vivo melanoma-targeting profiles when labeled with radiometals. Therefore, we hypothesized that ReCCMSH(Arg(11)) could be a good platform for the further development of an (18)F-labeled probe for PET of MC1R-positive malignant melanoma.In this study, the metallopeptide Ac-d-Lys-ReCCMSH(Arg(11)) was synthesized using conventional solid-phase peptide synthesis chemistry and a rhenium cyclization reaction. The resulting peptides were then labeled with N-succinimidyl-4-(18)F-fluorobenzoate ((18)F-SFB). The (18)F-labeled metallopeptides were further tested for their in vitro receptor binding affinities, in vivo biodistribution, and PET imaging properties.Both isomers of Ac-d-Lys-ReCCMSH(Arg(11)), named as RMSH-1 and RMSH-2, were purified and identified by high-performance liquid chromatography. The binding affinities of RMSH-1 and RMSH-2 and their respective (19)F-SFB-conjugated peptides ((19)F-FB-RMSH-1 and (19)F-FB-RMSH-2) were all determined to be within nanomolar range. Both (18)F-labeled metallopeptides showed good tumor uptake in the B16F10 murine model, with high MC1R expression, but much lower uptake in the A375M human melanoma xenografted in mice, indicating low MC1R expression. (18)F-FB-RMSH-1, when compared with (18)F-FB-RMSH-2, displayed more favorable in vivo performance in terms of slightly higher tumor uptakes and much lower accumulations in the kidney and liver at 2 h after injection. Small-animal PET of (18)F-FB-RMSH-1 and -2 in mice bearing B16F10 tumors at 1 and 2 h showed good tumor imaging quality. As expected, much lower tumor uptakes and poorer tumor-to-normal organ contrasts were observed for the A375M model than for the B16F10 model. (18)F-FB-RMSH-1 and -2 showed higher tumor uptake and better tumor retention than did (18)F-FB-NAPamide.Specific in vivo targeting of (18)F-FB-RMSH-1 to malignant melanoma was successfully achieved in preclinical models with high MC1R expression. Thus, the radiofluorinated metallopeptide (18)F-FB-RMSH-1 is a promising molecular probe for PET of MC1R-positive tumors.

  View details for DOI 10.2967/jnumed.109.062877

  View details for Web of Science ID 000272554100021

  View details for PubMedID 19837749

• Whole-body, real-time preclinical imaging of quantum dot fluorescence with time-gated detection JOURNAL OF BIOMEDICAL OPTICS May, A., Bhaumik, S., Gambhir, S. S., Zhan, C., Yazdanfar, S. 2009; 14 (6)

  Abstract

  We describe a wide-field preclinical imaging system optimized for time-gated detection of quantum dot fluorescence emission. As compared to continuous wave measurements, image contrast was substantially improved by suppression of short-lifetime background autofluorescence. Real-time (8 frames/s) biological imaging of subcutaneous quantum dot injections is demonstrated simultaneously in multiple living mice.

  View details for DOI 10.1117/1.3269675

  View details for Web of Science ID 000274267900004

  View details for PubMedID 20059235

  View details for PubMedCentralID PMC2801727

• Matrix-insensitive protein assays push the limits of biosensors in medicine NATURE MEDICINE Gaster, R. S., Hall, D. A., Nielsen, C. H., Osterfeld, S. J., Yu, H., Mach, K. E., Wilson, R. J., Murmann, B., Liao, J. C., Gambhir, S. S., Wang, S. X. 2009; 15 (11): 1327-U130

  Abstract

  Advances in biosensor technologies for in vitro diagnostics have the potential to transform the practice of medicine. Despite considerable work in the biosensor field, there is still no general sensing platform that can be ubiquitously applied to detect the constellation of biomolecules in diverse clinical samples (for example, serum, urine, cell lysates or saliva) with high sensitivity and large linear dynamic range. A major limitation confounding other technologies is signal distortion that occurs in various matrices due to heterogeneity in ionic strength, pH, temperature and autofluorescence. Here we present a magnetic nanosensor technology that is matrix insensitive yet still capable of rapid, multiplex protein detection with resolution down to attomolar concentrations and extensive linear dynamic range. The matrix insensitivity of our platform to various media demonstrates that our magnetic nanosensor technology can be directly applied to a variety of settings such as molecular biology, clinical diagnostics and biodefense.

  View details for DOI 10.1038/nm.2032

  View details for Web of Science ID 000271543700023

  View details for PubMedID 19820717

• A strategy for blood biomarker amplification and localization using ultrasound PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA D'Souza, A. L., Tseng, J. R., Pauly, K. B., Guccione, S., Rosenberg, J., Gambhir, S. S., Glazer, G. M. 2009; 106 (40): 17152-17157

  Abstract

  Blood biomarkers have significant potential applications in early detection and management of various diseases, including cancer. Most biomarkers are present in low concentrations in blood and are difficult to discriminate from noise. Furthermore, blood measurements of a biomarker do not provide information about the location(s) where it is produced. We hypothesize a previously undescribed strategy to increase the concentration of biomarkers in blood as well as localize the source of biomarker signal using ultrasound energy directly applied to tumor cells. We test and validate our hypothesis in cell culture experiments and mouse tumor xenograft models using the human colon cancer cell line LS174T, while measuring the biomarker carcinoembryonic antigen (CEA) before and after the use of ultrasound to liberate the biomarker from the tumor cells. The results demonstrate that the application of low-frequency ultrasound to tumor cells causes a significant release of tumor biomarker, which can be measured in the blood. Furthermore, we establish that this release is specific to the direct application of the ultrasound to the tumor, enabling a method for localization of biomarker production. This work shows that it is possible to use ultrasound to amplify and localize the source of CEA levels in blood of tumor-bearing mice and will allow for a previously undescribed way to determine the presence and localization of disease more accurately using a relatively simple and noninvasive strategy.

  View details for DOI 10.1073/pnas.0903437106

  View details for Web of Science ID 000270537500053

  View details for PubMedID 19805109

  View details for PubMedCentralID PMC2749842

• A novel multimodal approach to track neural progenitor cells in vivo 24th International Symposium on Cerebral Blood Flow and Metabolism/9th International Conference on Quantification of Brain Function with PET Pendharkar, A., De, A., Wang, H., Gaeta, X., Wang, N., Chua, J. Y., Andres, R., Chen, X., Gambhir, S. S., Guzman, R. NATURE PUBLISHING GROUP. 2009: S472–S473

  View details for Web of Science ID 000270329900575

• Creatine modulates survival, migration and differentiation in neural stem cells 24th International Symposium on Cerebral Blood Flow and Metabolism/9th International Conference on Quantification of Brain Function with PET Andres, R. H., Pendharkar, A., Guzman, R., De, A., Bliss, T. M., MacMillan, E., Svendsen, C., Gambhir, S. S., Widmer, H., Wallimann, T. NATURE PUBLISHING GROUP. 2009: S562–S562

  View details for Web of Science ID 000270329900691

• A Novel Estrogen Receptor Intramolecular Folding-based Titratable Transgene Expression System MOLECULAR THERAPY Paulmurugan, R., Padmanabhan, P., Ahn, B., Ray, S., Willmann, J. K., Massoud, T. F., Biswal, S., Gambhir, S. S. 2009; 17 (10): 1703-1711

  Abstract

  The use of regulated gene expression systems is important for successful gene therapy applications. In this study, ligand-induced structural change in the estrogen receptor (ER) was used to develop a novel ER intramolecular folding-based transcriptional activation system. The system was studied using ER-variants of different lengths, flanked on either side by the GAL4-DNA-binding domain and the VP16-transactivation domain (GAL4(DBD)-ER-VP16). The ER ligands of different types showed efficient ligand-regulated transactivation. We also characterized a bidirectional transactivation system based on the ER and demonstrated its utility in titrating both reporter and therapeutic gene expression. The ligand-regulated transactivation system developed by using a mutant form of the ER (G521T, lacking affinity for the endogenous ligand 17beta-estradiol, whereas maintaining affinity for other ligands) showed efficient activation by the ligand raloxifene in living mice without significant interference from the circulating endogenous ligand. The ligand-regulated transactivation system was used to test the therapeutic efficiency of the tumor suppressor protein p53 in HepG2 (p53(+/+)) and SKBr3 (p53(-/-)/mutant-p53(+/+)) cells in culture and tumor xenografts in living mice. The multifunctional capabilities of this system should be useful for gene therapy applications, to study ER biology, to evaluate gene regulation, ER ligand screening, and ER ligand biocharacterization in cells and living animals.

  View details for DOI 10.1038/mt.2009.171

  View details for PubMedID 19654568

• Melanin-Targeted Preclinical PET Imaging of Melanoma Metastasis JOURNAL OF NUCLEAR MEDICINE Ren, G., Miao, Z., Liu, H., Jiang, L., Limpa-Amara, N., Mahmood, A., Gambhir, S. S., Cheng, Z. 2009; 50 (10): 1692-1699

  Abstract

  Dialkylamino-alkyl-benzamides possess an affinity for melanin, suggesting that labeling of such benzamides with (18)F could potentially produce melanin-targeted PET probes able to identify melanotic melanoma metastases in vivo with high sensitivity and specificity.In this study, N-[2-(diethylamino)ethyl]-4-(18)F-fluorobenzamide ((18)F-FBZA) was synthesized via a 1-step conjugation reaction. The sigma-receptor binding affinity of (19)F-FBZA was determined along with the in vitro cellular uptake of radiofluorinated (18)F-FBZA in B16F10 cells. In vivo distribution and small-animal PET studies were conducted on mice bearing B16F10 melanoma, A375M amelanotic melanoma, and U87MG tumors, and comparative studies were performed with (18)F-FDG PET in the melanoma models.In vitro, uptake of (18)F-FBZA was significantly higher in B16F10 cells treated with l-tyrosine (P < 0.001). In vivo, (18)F-FBZA displayed significant tumor uptake; at 2 h, 5.94 +/- 1.83 percentage injected dose (%ID) per gram was observed in B16F10 tumors and only 0.75 +/- 0.09 %ID/g and 0.56 +/- 0.13 %ID/g was observed in amelanotic A375M and U87MG tumors, respectively. Lung uptake was significantly higher in murine lungs bearing melanotic B16F10 pulmonary metastases than in normal murine lungs (P < 0.01). Small-animal PET clearly identified melanotic lesions in both primary and pulmonary metastasis B16F10 tumor models. Coregistered micro-CT with small-animal PET along with biopsies further confirmed the presence of tumor lesions in the mouse lungs.(18)F-FBZA specifically targets primary and metastatic melanotic melanoma lesions with high tumor uptake and may have translational potential.

  View details for DOI 10.2967/jnumed.109.066175

  View details for Web of Science ID 000272553600023

  View details for PubMedID 19759116

• Prospective Evaluation of Tc-99m-MDP Scintigraphy, F-18 NaF PET/CT and F-18 FDG PET/CT for Detection of Skeletal Metastases Iagaru, A., Mittra, E., Dick, D., Gambhir, S. S. SPRINGER. 2009: S187–S187

  View details for Web of Science ID 000208690800141

• Efficacy of F-18-FDG PET/CT for Breast Cancer Mittra, E., Quon, A., Gambhir, S. S., Iagaru, A. SPRINGER. 2009: S176–S176

  View details for Web of Science ID 000208690800091

• Combined F-18 Fluoride and F-18 FDG PET/CT Scan for Evaluation of Malignancy Lagaru, A., Mittra, E., Dick, D., Quon, A., Goris, M. L., Gambhir, S. S. SPRINGER. 2009: S214–S214

  View details for Web of Science ID 000208690800268

• A 2-Helix Small Protein Labeled with Ga-68 for PET Imaging of HER2 Expression JOURNAL OF NUCLEAR MEDICINE Ren, G., Zhang, R., Liu, Z., Webster, J. M., Miao, Z., Gambhir, S. S., Syud, F. A., Cheng, Z. 2009; 50 (9): 1492-1499

  Abstract

  Affibody molecules are a class of scaffold proteins being developed into a generalizable approach to targeting tumors. Many 3-helix-based Affibody proteins have shown excellent in vivo properties for tumor imaging and therapy. By truncating one alpha-helix that is not responsible for receptor recognition in the Affibody and maturating the protein affinity through synthetic strategies, we have successfully identified in our previous research several small 2-helix proteins with excellent binding affinities to human epidermal growth factor receptor type 2 (HER2). With preferential properties such as faster blood clearance and tumor accumulation, lower immunogenic potential, and facile and economically viable synthetic schemes, we hypothesized that these 2-helix protein binders could become excellent molecular imaging probes for monitoring HER2 expression and modulation.In this study, a 2-helix small protein, MUT-DS, was chemically modified with a metal chelator, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). DOTA-MUT-DS was then site-specifically radiolabeled with an important PET radionuclide, (68)Ga. The resulting radiolabeled anti-HER2 2-helix molecule was further evaluated as a potential molecular probe for small-animal PET HER2 imaging in a SKOV3 tumor mouse model.The 2-helix DOTA-MUT-DS showed high HER2-binding affinity (dissociation constant, 4.76 nM). The radiolabeled probe displayed high stability in mouse serum and specificity toward HER2 in cell cultures. Biodistribution and small-animal PET studies further showed that (68)Ga-DOTA-MUT-DS had rapid and high SKOV3 tumor accumulation and quick clearance from normal organs. The specificity of (68)Ga-DOTA-MUT-DS for SKOV3 tumors was confirmed by monitoring modulation of HER2 protein on treatment of tumor mice with heat shock protein 90 inhibitor 17-N,N-dimethyl ethylene diamine-geldanamycin in vivo.This proof-of-concept research clearly demonstrated that synthetic 2-helix (68)Ga-DOTA-MUT-DS is a promising PET probe for imaging HER2 expression in vivo. The Affibody-derived small 2-helix protein scaffold has great potential for developing targeting agents for a variety of tumor-associated biomarkers.

  View details for DOI 10.2967/jnumed.109.064287

  View details for Web of Science ID 000272548900023

  View details for PubMedID 19690041

• Chemical tools for imaging glycosylation in vivo Chang, P. V., Prescher, J. A., Foss, C. A., Ray, P., Gambhir, S. S., Pomper, M. G., Bertozzi, C. R. AMER CHEMICAL SOC. 2009

  View details for Web of Science ID 000207861907424

• Visualizing Implanted Tumors in Mice with Magnetic Resonance Imaging Using Magnetotactic Bacteria CLINICAL CANCER RESEARCH Benoit, M. R., Mayer, D., Barak, Y., Chen, I. Y., Hu, W., Cheng, Z., Wang, S. X., Spielman, D. M., Gambhir, S. S., Matin, A. 2009; 15 (16): 5170-5177

  Abstract

  To determine if magnetotactic bacteria can target tumors in mice and provide positive contrast for visualization using magnetic resonance imaging.The ability of the magnetotactic bacterium, Magnetospirillum magneticum AMB-1 (referred to from here as AMB-1), to confer positive magnetic resonance imaging contrast was determined in vitro and in vivo. For the latter studies, AMB-1 were injected either i.t. or i.v. Bacterial growth conditions were manipulated to produce small (approximately 25-nm diameter) magnetite particles, which were observed using transmission electron microscopy. Tumor targeting was confirmed using 64Cu-labeled bacteria and positron emission tomography and by determination of viable cell counts recovered from different organs and the tumor.We show that AMB-1 bacteria with small magnetite particles generate T1-weighted positive contrast, enhancing in vivo visualization by magnetic resonance imaging. Following i.v. injection of 64Cu-labeled AMB-1, positron emission tomography imaging revealed increasing colonization of tumors and decreasing infection of organs after 4 hours. Viable cell counts showed that, by day 6, the bacteria had colonized tumors but were cleared completely from other organs. Magnetic resonance imaging showed a 1.22-fold (P = 0.003) increased positive contrast in tumors on day 2 and a 1.39-fold increase (P = 0.0007) on day 6.Magnetotactic bacteria can produce positive magnetic resonance imaging contrast and colonize mouse tumor xenografts, providing a potential tool for improved magnetic resonance imaging visualization in preclinical and translational studies to track cancer.

  View details for DOI 10.1158/1078-0432.CCR-08-3206

  View details for Web of Science ID 000269024900019

  View details for PubMedID 19671860

  View details for PubMedCentralID PMC3409839

• Multiplexed imaging of surface enhanced Raman scattering nanotags in living mice using noninvasive Raman spectroscopy PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Zavaleta, C. L., Smith, B. R., Walton, I., Doering, W., Davis, G., Shojaei, B., Natan, M. J., Gambhir, S. S. 2009; 106 (32): 13511-13516

  Abstract

  Raman spectroscopy is a newly developed, noninvasive preclinical imaging technique that offers picomolar sensitivity and multiplexing capabilities to the field of molecular imaging. In this study, we demonstrate the ability of Raman spectroscopy to separate the spectral fingerprints of up to 10 different types of surface enhanced Raman scattering (SERS) nanoparticles in a living mouse after s.c. injection. Based on these spectral results, we simultaneously injected the five most intense and spectrally unique SERS nanoparticles i.v. to image their natural accumulation in the liver. All five types of SERS nanoparticles were successfully identified and spectrally separated using our optimized noninvasive Raman imaging system. In addition, we were able to linearly correlate Raman signal with SERS concentration after injecting four spectrally unique SERS nanoparticles either s.c. (R(2) = 0.998) or i.v. (R(2) = 0.992). These results show great potential for multiplexed imaging in living subjects in cases in which several targeted SERS probes could offer better detection of multiple biomarkers associated with a specific disease.

  View details for DOI 10.1073/pnas.0813327106

  View details for Web of Science ID 000268877300066

  View details for PubMedID 19666578

  View details for PubMedCentralID PMC2726370

• Cys-diabody Quantum Dot Conjugates (ImmunoQdots) for Cancer Marker Detection BIOCONJUGATE CHEMISTRY Barat, B., Sirk, S. J., McCabe, K. E., Li, J., Lepin, E. J., Remenyi, R., Koh, A. L., Olafsen, T., Gambhir, S. S., Weiss, S., Wu, A. M. 2009; 20 (8): 1474-1481

  Abstract

  The present work demonstrates the use of small bivalent engineered antibody fragments, cys-diabodies, for biological modification of nanoscale particles such as quantum dots (Qdots) for detection of target antigens. Novel bioconjugated quantum dots known as immunoQdots (iQdots) were developed by thiol-specific oriented coupling of tumor specific cys-diabodies, at a position away from the antigen binding site to amino PEG CdSe/ZnS Qdots. Initially, amino PEG Qdot 655 were coupled with reduced anti-HER2 cys-diabody by amine-sulfhydryl-reactive linker [N-ε-maleimidocaproyloxy] succinimide ester (EMCS) to produce anti-HER2 iQdot 655. Spectral characterization of the conjugate revealed that the spectrum was symmetrical and essentially identical to unconjugated Qdot. Specific receptor binding activity of anti-HER2 iQdot 655 was confirmed by flow cytometry on HER2 positive and negative cells. Immunofluorescence results showed homogeneous surface labeling of the cell membrane with Qdot 655 conjugate. In addition, cys-diabodies specific for HER2, as well as prostate stem cell antigen (PSCA), were conjugated successfully with amino PEG Qdot 800. All of these iQdots retain the photoluminescence properties of the unconjugated Qdot 800 as well as the antigen binding specificity of the cys-diabody as demonstrated by flow cytometry. Simultaneous detection of two tumor antigens on LNCaP/PSCA prostate cancer cells (which express PSCA and HER2) in culture was possible using two iQdots, anti-HER2 iQdot 655 and anti-PSCA iQdot 800. Thus, these iQdots are potentially useful as optical probes for sensitive, multiplexed detection of surface markers on tumor cells. The present thiol-specific conjugation method demonstrates a general approach for site-specific oriented coupling of cys-diabodies to a wide variety of nanoparticles without disturbing the antigen binding site and maintaining small size compared to intact antibody.

  View details for DOI 10.1021/bc800421f

  View details for Web of Science ID 000269042100006

  View details for PubMedID 19642689

  View details for PubMedCentralID PMC2891877

• BRET3: a red-shifted bioluminescence resonance energy transfer (BRET)-based integrated platform for imaging protein-protein interactions from single live cells and living animals FASEB JOURNAL De, A., Ray, P., Loening, A. M., Gambhir, S. S. 2009; 23 (8): 2702-2709

  Abstract

  Taking advantage of the bioluminescence resonance energy transfer (BRET) phenomenon, we report the development of a highly photon-efficient, self-illuminating fusion protein combining a mutant red fluorescent protein (mOrange) and a mutant Renilla reniformis luciferase (RLuc8). This new BRET fusion protein (BRET3) exhibits severalfold improvement in light intensity in comparison with existing BRET fusion proteins. BRET3 also exhibits the most red-shifted light output (564-nm peak wavelength) of any reported bioluminescent protein that utilizes its natural substrate coelenterazine, a benefit of which is demonstrated at various tissue depths in small animals. The imaging utility of BRET3 at the single-cell level is demonstrated using an intramolecular sensor incorporating two mammalian target of rapamycin pathway proteins (FKBP12 and FRB) that dimerize only in the presence of rapamycin. With its increased photon intensity, red-shifted light output, and good spectral resolution (approximately 85 nm), BRET3 shows improved spatial and temporal resolution for measuring intracellular events in single cells and in living small animal models. The development of further BRET3-based assays will allow imaging of protein-protein interactions using a single assay directly scalable from intact living cells to small living subjects, allowing accelerated drug discovery.

  View details for DOI 10.1096/fj.08-118919

  View details for PubMedID 19351700

• Simulations of Virtual PET/CT 3-D Bronchoscopy Imaging Using a Physical Porcine Lung-Heart Phantom MOLECULAR IMAGING AND BIOLOGY Yerushalmi, D., Mullick, R., Quon, A., Fahrig, R., Pelc, N. J., Fann, J. I., Gambhir, S. S. 2009; 11 (4): 275-282

  Abstract

  We present a systematic approach for studying positron emission tomography-computed tomography (PET/CT) 3-D virtual fly-through endoscopy and for assessing the accuracy of this technology for visualizing and detecting endobronchial lesions as a function of focal lesion morphology and activity.Capsules designed to simulate endobronchial lesions were filled with activity and introduced into a porcine lung-heart phantom. PET/CT images were acquired, reconstructed, and volume rendered as 3-D fly-through and fly-around visualizations. Anatomical positioning of lesions seen on the 3-D-volume-rendered PET/CT images was compared to the actual position of the capsules.Lesion size was observed to be highly sensitive to PET threshold parameter settings and careful opacity and color transfer function parameter assignment.We have demonstrated a phantom model for studies of PET/CT 3-D virtual fly-through bronchoscopy and have applied this model for understanding the effect of PET thresholding on the visualization and detection of lesions.

  View details for DOI 10.1007/s11307-009-0201-8

  View details for Web of Science ID 000266830700010

  View details for PubMedID 19434462

• Imaging Gene Expression in Human Mesenchymal Stem Cells: From Small to Large Animals RADIOLOGY Willmann, J. K., Paulmurugan, R., Rodriguez-Porcel, M., Stein, W., Brinton, T. J., Connolly, A. J., Nielsen, C. H., Lutz, A. M., Lyons, J., Ikeno, F., Suzuki, Y., Rosenberg, J., Chen, I. Y., Wu, J. C., Yeung, A. C., Yock, P., Robbins, R. C., Gambhir, S. S. 2009; 252 (1): 117-127

  Abstract

  To evaluate the feasibility of reporter gene imaging in implanted human mesenchymal stem cells (MSCs) in porcine myocardium by using clinical positron emission tomography (PET)-computed tomography (CT) scanning.Animal protocols were approved by the Institutional Administrative Panel on Laboratory Animal Care. Transduction of human MSCs by using different doses of adenovirus that contained a cytomegalovirus (CMV) promoter driving the mutant herpes simplex virus type 1 thymidine kinase reporter gene (Ad-CMV-HSV1-sr39tk) was characterized in a cell culture. A total of 2.25 x 10(6) transduced (n = 5) and control nontransduced (n = 5) human MSCs were injected into the myocardium of 10 rats, and reporter gene expression in human MSCs was visualized with micro-PET by using the radiotracer 9-(4-[fluorine 18]-fluoro-3-hydroxymethylbutyl)-guanine (FHBG). Different numbers of transduced human MSCs suspended in either phosphate-buffered saline (PBS) (n = 4) or matrigel (n = 5) were injected into the myocardium of nine swine, and gene expression was visualized with a clinical PET-CT. For analysis of cell culture experiments, linear regression analyses combined with a t test were performed. To test differences in radiotracer uptake between injected and remote myocardium in both rats and swine, one-sided paired Wilcoxon tests were performed. In swine experiments, a linear regression of radiotracer uptake ratio on the number of injected transduced human MSCs was performed.In cell culture, there was a viral dose-dependent increase of gene expression and FHBG accumulation in human MSCs. Human MSC viability was 96.7% (multiplicity of infection, 250). Cardiac FHBG uptake in rats was significantly elevated (P < .0001) after human MSC injection (0.0054% injected dose [ID]/g +/- 0.0007 [standard deviation]) compared with that in the remote myocardium (0.0003% ID/g +/- 0.0001). In swine, myocardial radiotracer uptake was not elevated after injection of up to 100 x 10(6) human MSCs (PBS group). In the matrigel group, signal-to-background ratio increased to 1.87 after injection of 100 x 10(6) human MSCs and positively correlated (R(2) = 0.97, P < .001) with the number of administered human MSCs.Reporter gene imaging in human MSCs can be translated to large animals. The study highlights the importance of co-administering a "scaffold" for increasing intramyocardial retention of human MSCs.

  View details for DOI 10.1148/radiol.2513081616

  View details for Web of Science ID 000268362900015

  View details for PubMedID 19366903

  View details for PubMedCentralID PMC2702468

• Role of Oxidative Stress in Stem Cell Survival 10th Annual Conference on Arteriosclerosis, Thrombosis and Vascular Biology Peterson, K. M., Abdelrhaman, A., Gambhir, S. S., Rodriguez-Porcel, M. G. LIPPINCOTT WILLIAMS & WILKINS. 2009: E88–E88

  View details for Web of Science ID 000267102500416

• Lymphoid tissue-specific homing of bone marrow-derived dendritic cells BLOOD Creusot, R. J., Yaghoubi, S. S., Chang, P., Chia, J., Contag, C. H., Gambhir, S. S., Fathman, C. G. 2009; 113 (26): 6638-6647

  Abstract

  Because of their potent immunoregulatory capacity, dendritic cells (DCs) have been exploited as therapeutic tools to boost immune responses against tumors or pathogens, or dampen autoimmune or allergic responses. Murine bone marrow-derived DCs (BM-DCs) are the closest known equivalent of the blood monocyte-derived DCs that have been used for human therapy. Current imaging methods have proven unable to properly address the migration of injected DCs to small and deep tissues in mice and humans. This study presents the first extensive analysis of BM-DC homing to lymph nodes (and other selected tissues) after intravenous and intraperitoneal inoculation. After intravenous delivery, DCs accumulated in the spleen, and preferentially in the pancreatic and lung-draining lymph nodes. In contrast, DCs injected intraperitoneally were found predominantly in peritoneal lymph nodes (pancreatic in particular), and in omentum-associated lymphoid tissue. This uneven distribution of BM-DCs, independent of the mouse strain and also observed within pancreatic lymph nodes, resulted in the uneven induction of immune response in different lymphoid tissues. These data have important implications for the design of systemic cellular therapy with DCs, and in particular underlie a previously unsuspected potential for specific treatment of diseases such as autoimmune diabetes and pancreatic cancer.

  View details for DOI 10.1182/blood-2009-02-204321

  View details for Web of Science ID 000267789600024

  View details for PubMedID 19363220

  View details for PubMedCentralID PMC2710920

• OXIDATIVE STRESS BLOCKADE IMPROVES STEM CELL SURVIVAL Peterson, K., Aly, A., Gambhir, S., Rodriguez-Porcel, M. ELSEVIER IRELAND LTD. 2009

  View details for Web of Science ID 000207957101474

• Tumor metabolic phenotypes on 18F FDG PET. Journal of nuclear medicine : official publication, Society of Nuclear Medicine Wong, C. O., Khong, P. L. 2009; 50 (6): 1010-1; author reply 1011-12

  View details for DOI 10.2967/jnumed.108.061010

  View details for PubMedID 19443584

• Tumor Metabolic Phenotypes on F-18 FDG PET REPLY JOURNAL OF NUCLEAR MEDICINE Iagaru, A. H., Gambhir, S. S., Goris, M. L. 2009; 50 (6): 1011-1012

  View details for DOI 10.2967/jnumed.109.061945

  View details for Web of Science ID 000272488000034

• Engineered Two-Helix Small Proteins for Molecular Recognition CHEMBIOCHEM Webster, J. M., Zhang, R., Gambhir, S. S., Cheng, Z., Syud, F. A. 2009; 10 (8): 1293-1296

  Abstract

  Less is more: By starting with a high-affinity HER2-binding 3-helix affibody molecule, we successfully developed 2-helix small protein binders with 5 nM affinities by using a combination of several different strategies. Our efforts clearly suggest that 2-helix small proteins against important tumor targets can be obtained by rational protein design and engineering.

  View details for DOI 10.1002/cbic.200900062

  View details for Web of Science ID 000266561500003

  View details for PubMedID 19422008

• Comparison of Optical Bioluminescence Reporter Gene and Superparamagnetic Iron Oxide MR Contrast Agent as Cell Markers for Noninvasive Imaging of Cardiac Cell Transplantation MOLECULAR IMAGING AND BIOLOGY Chen, I. Y., Greve, J. M., Gheysens, O., Willmann, J. K., Rodriguez-Porcel, M., Chu, P., Sheikh, A. Y., Faranesh, A. Z., Paulmurugan, R., Yang, P. C., Wu, J. C., Gambhir, S. S. 2009; 11 (3): 178-187

  Abstract

  In this study, we compared firefly luciferase (Fluc) reporter gene and superparamagnetic iron oxide (Feridex) as cell markers for longitudinal monitoring of cardiomyoblast graft survival using optical bioluminescence imaging (BLI) and magnetic resonance imaging (MRI), respectively.Rats (n = 31) underwent an intramyocardial injection of cardiomyoblasts (2 x 10(6)) labeled with Fluc, Feridex, or no marker (control) or an injection of Feridex alone (75 microg). Afterward, rats were serially imaged with BLI or MRI and killed at different time points for histological analysis.BLI revealed a drastically different cell survival kinetics (half-life = 2.65 days over 6 days) than that revealed by MRI (half-life = 16.8 days over 80 days). Injection of Feridex alone led to prolonged tissue retention of Feridex (> or =16 days) and persistent MR signal (> or =42 days).Fluc BLI reporter gene imaging is a more accurate gauge of transplanted cell survival as compared to MRI of Feridex-labeled cells.

  View details for DOI 10.1007/s11307-008-0182-z

  View details for PubMedID 19034584

• A protein scaffold based molecule for EGFR PET imaging Miao, Z., Ren, G., Liu, H., Jiang, L., Cheng, Z., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2009

  View details for DOI 10.2967/jnumed.109.062877

  View details for Web of Science ID 000444145700385

• CT and FDG PET/CT evaluation of response to NV1020 for liver colorectal metastases Iagaru, A., Sze, D., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2009

  View details for Web of Science ID 000444145700287

• Melanin targeted molecular imaging of melanoma metastasis using a 18F-labeled benzamide analog Ren, G., Miao, Z., Liu, H., Jiang, L., Limpa-Amara, N., Mahmood, A., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2009

  View details for Web of Science ID 000444145700333

• Molecular imaging of malignant melanoma using a 18F-labeled metallopeptide Ren, G., Liu, Z., Miao, Z., Liu, H., Jiang, L., Subbarayan, M., Chin, F., Zhang, L., Gambhir, S., Cheng, Z. SOC NUCLEAR MEDICINE INC. 2009

  View details for Web of Science ID 000444145701587

• Value of FDG PET/CT for the restaging of colorectal cancer after treatment with Erbitux. Mittra, E., Iagaru, A., Kunz, P., Quon, A., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2009

  View details for Web of Science ID 000444145702017

• A novel strategy for a cocktail 18F fluoride and 18F FDG PET/CT scan for evaluation of malignancy: Results of the pilot phase study Iagaru, A., Mittra, E., Quon, A., Goris, M., Gambhir, S. SOC NUCLEAR MEDICINE INC. 2009

  View details for Web of Science ID 000444145700133

• Optimized imaging of the growth and metastasis of human breast cancer stem cells in immunodeficient mice Liu, H., Patel, M., Prescher, J., Qian, D., Dalerba, P., Lin, J., Shimono, Y., Dirbas, F., Contag, C., Gambhir, S., Clarke, M. AMER ASSOC CANCER RESEARCH. 2009

  View details for Web of Science ID 000209702603139

• A Potent, Imaging Adenoviral Vector Driven by the Cancer-selective Mucin-1 Promoter That Targets Breast Cancer Metastasis CLINICAL CANCER RESEARCH Huyn, S. T., Burton, J. B., Sato, M., Carey, M., Gambhir, S. S., Wu, L. 2009; 15 (9): 3126-3134

  Abstract

  With breast cancer, early detection and proper staging are critical, and will often influence both the treatment regimen and the therapeutic outcome for those affected with this disease. Improvements in these areas will play a profound role in reducing mortality from breast cancer.In this work we developed a breast cancer-targeted serotype 5 adenoviral vector, utilizing the tumor-specific mucin-1 promoter in combination with the two-step transcriptional amplification system, a system used to augment the activity of weak tissue-specific promoters.We showed the strong specificity of this tumor-selective adenovirus to express the luciferase optical imaging gene, leading to diagnostic signals that enabled detection of sentinel lymph node metastasis of breast cancer. Furthermore, we were able to target hepatic metastases following systemic administration of this mucin-1 selective virus.Collectively, we showed that the amplified mucin-1 promoter-driven vector is able to deliver to and selectively express a desirable transgene in metastatic lesions of breast tumors. This work has strong clinical relevance to current diagnostic staging approaches, and could add to targeted therapeutic strategies to advance the fight against breast cancer.

  View details for DOI 10.1158/1078-0432.CCR-08-2666

  View details for Web of Science ID 000265712100022

  View details for PubMedID 19366829

  View details for PubMedCentralID PMC2830790

• Molecular Imaging of Phosphorylation Events for Drug Development MOLECULAR IMAGING AND BIOLOGY CHAN, C. T., Paulmurugan, R., Reeves, R. E., Solow-Cordero, D., Gambhir, S. S. 2009; 11 (3): 144-158

  Abstract

  Protein phosphorylation mediated by protein kinases controls numerous cellular processes. A genetically encoded, generalizable split firefly luciferase (FL)-assisted complementation system was developed for noninvasive monitoring phosphorylation events and efficacies of kinase inhibitors in cell culture and in small living subjects by optical bioluminescence imaging.An Akt sensor (AST) was constructed to monitor Akt phosphorylation and the effect of different PI-3K and Akt inhibitors. Specificity of AST was determined using a non-phosphorylable mutant sensor containing an alanine substitution (ASA).The PI-3K inhibitor LY294002 and Akt kinase inhibitor perifosine led to temporal- and dose-dependent increases in complemented FL activities in 293T human kidney cancer cells stably expressing AST (293T/AST) but not in 293T/ASA cells. Inhibition of endogenous Akt phosphorylation and kinase activities by perifosine also correlated with increase in complemented FL activities in 293T/AST cells but not in 293T/ASA cells. Treatment of nude mice bearing 293T/AST xenografts with perifosine led to a 2-fold increase in complemented FL activities compared to that of 293T/ASA xenografts. Our system was used to screen a small chemical library for novel modulators of Akt kinase activity.This generalizable approach for noninvasive monitoring of phosphorylation events will accelerate the discovery and validation of novel kinase inhibitors and modulators of phosphorylation events.

  View details for DOI 10.1007/s11307-008-0187-7

  View details for Web of Science ID 000265686900002

  View details for PubMedID 19048345

  View details for PubMedCentralID PMC4154800

• Novel Strategy for a Cocktail F-18-Fluoride and F-18-FDG PET/CT Scan for Evaluation of Malignancy: Results of the Pilot-Phase Study JOURNAL OF NUCLEAR MEDICINE Iagaru, A., Mittra, E., Yaghoubi, S. S., Dick, D. W., Quon, A., Goris, M. L., Gambhir, S. S. 2009; 50 (4): 501-505

  Abstract

  (18)F-FDG PET/CT is used for detecting cancer and monitoring cancer response to therapy. However, because of the variable rates of glucose metabolism, not all cancers are identified reliably. Sodium (18)F was previously used for bone imaging and can be used as a PET/CT skeletal tracer. The combined administration of (18)F and (18)F-FDG in a single PET/CT study for cancer detection has not been reported to date.This is a prospective pilot study (November 2007-November 2008) of 14 patients with proven malignancy (6 sarcoma, 3 prostate cancer, 2 breast cancer, 1 colon cancer, 1 lung cancer, and 1 malignant paraganglioma) who underwent separate (18)F PET/CT and (18)F-FDG PET/CT and combined (18)F/(18)F-FDG PET/CT scans for the evaluation of malignancy (a total of 3 scans each). There were 11 men and 3 women (age range, 19-75 y; average, 50.4 y).Interpretation of the combined (18)F/(18)F-FDG PET/CT scans compared favorably with that of the (18)F-FDG PET/CT (no lesions missed) and the (18)F PET/CT scans (only 1 skull lesion seen on an (18)F PET/CT scan was missed on the corresponding combined scan). Through image processing, the combined (18)F/(18)F-FDG scan yielded results for bone radiotracer uptake comparable to those of the (18)F PET/CT scan performed separately.Our pilot-phase prospective trial demonstrates that the combined (18)F/(18)F-FDG administration followed by a single PET/CT scan is feasible for cancer detection. This combined method opens the possibility for improved patient care and reduction in health care costs.

  View details for DOI 10.2967/jnumed.108.058339

  View details for Web of Science ID 000272487200003

  View details for PubMedID 19289439

• Human adipose tissue-derived mesenchymal stromal cells as vehicles for tumor bystander effect: a model based on bioluminescence imaging GENE THERAPY Vilalta, M., Degano, I. R., Bago, J., Aguilar, E., Gambhir, S. S., Rubio, N., Blanco, J. 2009; 16 (4): 547-557

  Abstract

  Human adipose tissue mesenchymal stromal cells (AMSCs) share common traits, including similar differentiation potential and cell surface markers, with their bone marrow counterparts. Owing to their general availability, higher abundance and ease of isolation AMSCs may be convenient autologous delivery vehicles for localized tumor therapy. We demonstrate a model for tumor therapy development based on the use of AMSCs expressing renilla luciferase and thymidine kinase, as cellular vehicles for ganciclovir-mediated bystander killing of firefly luciferase expressing tumors, and noninvasive bioluminescence imaging to continuously monitor both, tumor cells and AMSCs. We show that the therapy delivering AMSCs survive long time within tumors, optimize the ratio of AMSCs to tumor cells for therapy, and asses the therapeutic effect in real time. Treatment of mice bearing prostate tumors plus therapeutic AMSCs with the prodrug ganciclovir induced bystander killing effect, reducing the number of tumor cells to 1.5 % that of control tumors. Thus, AMSCs could be useful vehicles to deliver localized therapy, with potential for clinical application in inoperable tumors and surgical borders after tumor resection. This approach, useful to evaluate efficiency of therapeutic models, should facilitate the selection of cell types, dosages, therapeutic agents and treatment protocols for cell-based therapies of specific tumors.

  View details for DOI 10.1038/gt.2008.176

  View details for Web of Science ID 000265021400012

  View details for PubMedID 19092860

• A Novel High-Sensitivity Rapid-Acquisition Single-Photon Cardiac Imaging Camera JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Berman, D. S., Ziffer, J., Nagler, M., Sandler, M., Patton, J., Hutton, B., Sharir, T., Ben Haim, S., Ben Haim, S. 2009; 50 (4): 635-643

  Abstract

  This study described and validated a new solid-state single-photon gamma-camera and compared it with a conventional-SPECT Anger camera. The compact new camera uses a unique method for localizing gamma-photon information with a bank of 9 solid-state detector columns with tungsten collimators that rotate independently.Several phantom studies were performed comparing the new technology with conventional-SPECT technology. These included measurements of line sources and single- and dual-radionuclide studies of a torso phantom. Simulations were also performed using a cardiothoracic phantom. Furthermore, 18 patients were scanned with both the new camera and a conventional-SPECT camera.The new camera had a count sensitivity that was 10 times higher than that of the conventional camera and a compensated spatial resolution that was moderately better. Dual-radionuclide studies using a phantom show the further potential of the new camera for a 2-tracer simultaneous acquisition. Two-minute clinical studies with the new camera and 11-min studies with the conventional camera qualitatively showed good-to-excellent image quality and improved myocardial edge definition for the new camera.These initial performance characteristics of a new solid-state single-photon gamma-camera offer great promise for clinical dynamic SPECT protocols, with important implications for applications in nuclear cardiology and molecular imaging.

  View details for DOI 10.2967/jnumed.108.060020

  View details for Web of Science ID 000272487200021

  View details for PubMedID 19339672

• A NEAR-INFRARED FLUORESCENT DEOXYGLUCOSE DERIVATIVE FOR OPTICAL IMAGING OF EXPERIMENTAL ARTHRITIS JOURNAL OF INNOVATIVE OPTICAL HEALTH SCIENCES Liu, X., Xiong, Z., Lee, S., Levi, J., Yaghoubi, S., Biswal, S., Gambhir, S. S., Cheng, Z. 2009; 2 (2): 179-187

  View details for DOI 10.1142/S1793545809000450

  View details for Web of Science ID 000208647200008

• Engineered Knottin Peptides: A New Class of Agents for Imaging Integrin Expression in Living Subjects CANCER RESEARCH Kimura, R. H., Cheng, Z., Gambhir, S. S., Cochran, J. R. 2009; 69 (6): 2435-2442

  Abstract

  There is a critical need for molecular imaging agents to detect cell surface integrin receptors that are present in human cancers. Previously, we used directed evolution to engineer knottin peptides that bind with high affinity ( approximately 10 to 30 nmol/L) to integrin receptors that are overexpressed on the surface of tumor cells and the tumor neovasculature. To evaluate these peptides as molecular imaging agents, we site-specifically conjugated Cy5.5 or (64)Cu-1,4,7,10-tetra-azacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) to their N termini, and used optical and positron emission tomography (PET) imaging to measure their uptake and biodistribution in U87MG glioblastoma murine xenograft models. NIR fluorescence and microPET imaging both showed that integrin binding affinity plays a strong role in the tumor uptake of knottin peptides. Tumor uptake at 1 hour postinjection for two high-affinity (IC(50), approximately 20 nmol/L) (64)Cu-DOTA-conjugated knottin peptides was 4.47% +/- 1.21% and 4.56% +/- 0.64% injected dose/gram (%ID/g), compared with a low-affinity knottin peptide (IC(50), approximately 0.4 mumol/L; 1.48 +/- 0.53%ID/g) and c(RGDyK) (IC(50), approximately 1 mumol/L; 2.32 +/- 0.55%ID/g), a low-affinity cyclic pentapeptide under clinical development. Furthermore, (64)Cu-DOTA-conjugated knottin peptides generated lower levels of nonspecific liver uptake ( approximately 2%ID/g) compared with c(RGDyK) ( approximately 4%ID/g) 1 hour postinjection. MicroPET imaging results were confirmed by in vivo biodistribution studies. (64)Cu-DOTA-conjugated knottin peptides were stable in mouse serum, and in vivo metabolite analysis showed minimal degradation in the blood or tumor upon injection. Thus, engineered integrin-binding knottin peptides show great potential as clinical diagnostics for a variety of cancers.

  View details for DOI 10.1158/0008-5472.CAN-08-2495

  View details for Web of Science ID 000264541300037

  View details for PubMedID 19276378

  View details for PubMedCentralID PMC2833353

• Stem Cell-Mediated Accelerated Bone Healing Observed with in Vivo Molecular and Small Animal Imaging Technologies in a Model of Skeletal Injury JOURNAL OF ORTHOPAEDIC RESEARCH Lee, S., Padmanabhan, P., Ray, P., Gambhir, S. S., Doyle, T., Contag, C., Goodman, S. B., Biswal, S. 2009; 27 (3): 295-302

  Abstract

  Adult stem cells are promising therapeutic reagents for skeletal regeneration. We hope to validate by molecular imaging technologies the in vivo life cycle of adipose-derived multipotent cells (ADMCs) in an animal model of skeletal injury. Primary ADMCs were lentivirally transfected with a fusion reporter gene and injected intravenously into mice with bone injury or sham operation. Bioluminescence imaging (BLI), [(18)F]FHBG (9-(fluoro-hydroxy-methyl-butyl-guanine)-micro-PET, [(18)F]Fluoride ion micro-PET and micro-CT were performed to monitor stem cells and their effect. Bioluminescence microscopy and immunohistochemistry were done for histological confirmation. BLI showed ADMC's traffic from the lungs then to the injury site. BLI microscopy and immunohistochemistry confirmed the ADMCs in the bone defect. Micro-CT measurements showed increased bone healing in the cell-injected group compared to the noninjected group at postoperative day 7 (p < 0.05). Systemically administered ADMC's traffic to the site of skeletal injury and facilitate bone healing, as demonstrated by molecular and small animal imaging. Molecular imaging technologies can validate the usage of adult adipose tissue-derived multipotent cells to promote fracture healing. Imaging can in the future help establish therapeutic strategies including dosage and administration route.

  View details for DOI 10.1002/jor.20736

  View details for PubMedID 18752273

• A Novel Method for Direct Site-Specific Radiolabeling of Peptides Using [F-18]FDG BIOCONJUGATE CHEMISTRY Namavari, M., Cheng, Z., Zhang, R., De, A., Levi, J., Hoerner, J. K., Yaghoubi, S. S., Syud, F. A., Gambhir, S. S. 2009; 20 (3): 432-436

  Abstract

  We have used the well-accepted and easily available 2-[(18)F]fluoro-2-deoxyglucose ([(18)F]FDG) positron emission tomography (PET) tracer as a prosthetic group for synthesis of (18)F-labeled peptides. We herein report the synthesis of [(18)F]FDG-RGD ((18)F labeled linear RGD) and [(18)F]FDG-cyclo(RGD(D)YK) ((18)F labeled cyclic RGD) as examples of the use of [(18)F]FDG. We have successfully prepared [(18)F]FDG-RGD and [(18)F]FDG-cyclo(RGD(D)YK) in 27.5% and 41% radiochemical yields (decay corrected) respectively. The receptor binding affinity study of FDG-cyclo(RGD(D)YK) for integrin alpha(v)beta(3), using alpha(v)beta(3) positive U87MG cells confirmed a competitive displacement with (125)I-echistatin as a radioligand. The IC(50) value for FDG-cyclo(RGD(D)YK) was determined to be 0.67 +/- 0.19 muM. High-contrast small animal PET images with relatively moderate tumor uptake were observed for [(18)F]FDG-RGD and [(18)F]FDG-cyclo(RGD(D)YK) as PET probes in xenograft models expressing alpha(v)beta(3) integrin. In conclusion, we have successfully used [(18)F]FDG as a prosthetic group to prepare (18)F]FDG-RGD and [(18)F]FDG-cyclic[RGD(D)YK] based on a simple one-step radiosynthesis. The one-step radiosynthesis methodology consists of chemoselective oxime formation between an aminooxy-functionalized peptide and [(18)F]FDG. The results have implications for radiolabeling of other macromolecules and would lead to a very simple strategy for routine preclinical and clinical use.

  View details for DOI 10.1021/bc800422b

  View details for Web of Science ID 000264389800005

  View details for PubMedID 19226160

  View details for PubMedCentralID PMC2765576

• Development of a breast cancer brain metastases model to study I-131 radioablative therapy. 31st Annual Meeting of the San Antonio Breast Cancer Symposium Renier, C., De, A., Hou, L., Dunkel, J., Sun, A., Prugpichailers, T., Gambhir, S. S., Tse, V., Wapnir, I. L. AMER ASSOC CANCER RESEARCH. 2009: 159S–160S

  View details for Web of Science ID 000262583200304

• Multimodality Molecular Imaging of Transplanted Neural Progenitor Cells 16th Annual Meeting of the American-Society-for-Neural-Therapy-and-Repair PENDHARKAR, A. V., De, A., Wang, H., Gaeta, X., Wang, N., Andres, R. H., Chen, X., Gambhir, S. S., Guzman, R. COGNIZANT COMMUNICATION CORP. 2009: 230–31

  View details for Web of Science ID 000266340800077

• Combined 18F-FDG anf Fluoride Approach in PET/CT Imaging: Is There a Clinical Future? Journal of Nuclear Medicine Iagaru, A., Mittra, E., Goris, E., Gambhir, S. S. 2009
• Human adipose tissue-derived menesenchymal stromal cells as vehicles for tumor bystander effect: A model based on bioluminescence imaging Gene Therapy Vilalta M, Degano I, Bago J, Aguilar E, Gambhir SS, Rubio N, Blanco J 2009; 16 (4): 547-557
• Effects of Creatine on Survival, Migration, and Differentiation of Neural Stem Cells 16th Annual Meeting of the American-Society-for-Neural-Therapy-and-Repair Andres, R. H., PENDHARKAR, A. V., Guzman, R., De, A., Bliss, T. M., McMillan, E., Svendsen, C. N., Gambhir, S. S., Widmer, H. R., Wallimann, T., Steinberg, G. K. COGNIZANT COMMUNICATION CORP. 2009: 208–

  View details for Web of Science ID 000266340800014

• Controlling the selection stringency of phage display using a microfluidic device LAB ON A CHIP Liu, Y., Adams, J. D., Turner, K., Cochran, F. V., Gambhir, S. S., Soh, H. T. 2009; 9 (8): 1033-1036

  Abstract

  We report the utilization of microfluidic technology to phage selection and demonstrate that accurate control of washing stringency in our microfluidic magnetic separator (MMS) directly impacts the diversity of isolated peptide sequences. Reproducible generation of magnetic and fluidic forces allows controlled washing conditions that enable rapid convergence of selected peptide sequences. These findings may provide a foundation for the development of automated microsystems for rapid in vitro directed evolution of affinity reagents.

  View details for DOI 10.1039/b820985e

  View details for Web of Science ID 000264978800001

  View details for PubMedID 19350081

• Endogenous NIS expression in triple negative breast cancers Annals of Surgical Oncology Renier C, Yao C, Goris M, Ghosh M, Katznelson L, Nowles K, Gambhir SS, Wapnir I 2009; 16 (4): 962-968
• Lymphoid tissue-specific homing of bone marrow-derived dendritic cells Blood Creusot R, Yaghoubi S, Chang P, Chia J, Contag C, Gambhir SS, Fathman C 2009; 113 (26): 6638-47
• Embryonic stem cell-derived endothelial cells for treatment of hindlimb ischemia. Journal of visualized experiments : JoVE Huang, N. F., Niiyama, H., De, A., Gambhir, S. S., Cooke, J. P. 2009

  Abstract

  Peripheral arterial disease (PAD) results from narrowing of the peripheral arteries that supply oxygenated blood and nutrients to the legs and feet, This pathology causes symptoms such as intermittent claudication (pain with walking), painful ischemic ulcerations, or even limb-threatening gangrene. It is generally believed that the vascular endothelium, a monolayer of endothelial cells that invests the luminal surface of all blood and lymphatic vessels, plays a dominant role in vascular homeostasis and vascular regeneration. As a result, stem cell-based regeneration of the endothelium may be a promising approach for treating PAD. In this video, we demonstrate the transplantation of embryonic stem cell (ESC)-derived endothelial cells for treatment of unilateral hindimb ischemia as a model of PAD, followed by non-invasive tracking of cell homing and survival by bioluminescence imaging. The specific materials and procedures for cell delivery and imaging will be described. This protocol follows another publication in describing the induction of hindlimb ischemia by Niiyama et al.

  View details for DOI 10.3791/1034

  View details for PubMedID 19229180

  View details for PubMedCentralID PMC2781824

• Noninvasive detection of therapeutic cytolytic T cells with F-18-FHBG PET in a patient with glioma NATURE CLINICAL PRACTICE ONCOLOGY Yaghoubi, S. S., Jensen, M. C., Satyamurthy, N., Budhiraja, S., Paik, D., Czernin, J., Gambhir, S. S. 2009; 6 (1): 53-58

  Abstract

  A 57-year-old man had been diagnosed with grade IV glioblastoma multiforme and was enrolled in a trial of adoptive cellular immunotherapy. The trial involved infusion of ex vivo expanded autologous cytolytic CD8+ T cells (CTLs), genetically engineered to express the interleukin 13 zetakine gene (which encodes a receptor protein that targets these T cells to tumor cells) and the herpes simplex virus 1 thymidine kinase (HSV1 tk) suicide gene, and PET imaging reporter gene.MRI, whole-body and brain PET scan with (18)F-radiolabelled 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine ((18)F-FHBG) to detect CTLs that express HSV1 tk, and safety monitoring after injection of (18)F-FHBG.MRI detected grade III-IV glioblastoma multiforme plus two tumors recurrences that developed after resection of the initial tumor.Surgical resection of primary glioblastoma tumor, enrollment in CTL therapy trial, reresection of glioma recurrences, infusion of approximately 1 x 10(9) CTLs into the site of tumor reresection, and (18)F-FHBG PET scan to detect infused CTLs.

  View details for DOI 10.1038/ncponc1278

  View details for Web of Science ID 000261845300011

  View details for PubMedID 19015650

  View details for PubMedCentralID PMC3526373

• A novel high-sensitivity rapid acquisition single photon cardiac imaging camera Journal of Nuclear Medicine Gambhir SS, Berman D, Ziffer J, Nagler M, Sandler M, Patton J, Hutton B, Sharir T, Haim S 2009; 50 (4): 635-643
• An Engineered Knottin Peptide Labeled with [18F] for PET Imaging of Integrin Expression. Bioconjugate Chemistry Miao Z, Ren G, Liu H, Kimura R, Jiang L, Cochran J, Gambhir SS, Cheng Z. 2009; 20(12): 2342-2347
• Trafficking Mesenchymal Stem Cell Engraftment and Differentiation in Tumor-Bearing Mice by Bioluminescence Imaging STEM CELLS Wang, H., Cao, F., De, A., Cao, Y., Contag, C., Gambhir, S. S., Wu, J. C., Chen, X. 2009; 27 (7): 1548-1558

  Abstract

  The objective of the study was to track the distribution and differentiation of mesenchymal stem cells (MSCs) in tumor-bearing mice. The 4T1 murine breast cancer cells were labeled with renilla luciferase-monomeric red fluorescence protein (rLuc-mRFP) reporter gene. The MSCs labeled with firefly luciferase-enhanced green fluorescence protein (fLuc-eGFP) reporter gene (MSCs-R) were isolated from L2G85 transgenic mice that constitutively express fLuc-eGFP reporter gene. To study the tumor tropism of MSCs, we established both subcutaneous and lung metastasis models. In lung metastasis tumor mice, we injected MSCs-R intravenously either on the same day or 4 days after 4T1 tumor cell injection. In subcutaneous tumor mice, we injected MSCs-R intravenously 7 days after subcutaneous 4T1 tumor inoculation. The tumor growth was monitored by rLuc bioluminescence imaging (BLI). The fate of MSCs-R was monitored by fLuc BLI. The localization of MSCs-R in tumors was examined histologically. The osteogenic and adipogenic differentiation of MSCs-R was investigated by alizarin red S and oil red O staining, respectively. The mechanism of the dissimilar differentiation potential of MSCs-R under different tumor microenvironments was investigated. We found that the 4T1 cells were successfully labeled with rLuc-mRFP. The MSCs-R isolated from L2G85 transgenic mice constitutively express fLuc-eGFP reporter gene. When injected intravenously, MSCs-R survived, proliferated, and differentiated in tumor sites but not elsewhere. The localization of GFP(+) MSCs-R in tumor lesions was confirmed ex vivo. In conclusion, the MSCs-R can selectively localize, survive, and proliferate in both subcutaneous tumor and lung metastasis as evidenced by noninvasive bioluminescence imaging and ex vivo validation. The MSCs-R migrated to lung tumor differentiated into osteoblasts, whereas the MSCs-R targeting subcutaneous tumor differentiated into adipocytes.

  View details for DOI 10.1002/stem.81

  View details for PubMedID 19544460

• AUTOMATED RADIOSYNTHESIS OF [F-18]EF-5 FOR IMAGING HYPOXIA IN HUMAN Chin, F. T., Subbarayan, M., Sorger, J., Gambhir, S. S., Graves, E. E. WILEY-BLACKWELL. 2009: S274–S274

  View details for Web of Science ID 000268724900275

• Particle Size, Surface Coating, and PEGylation Influence the Biodistribution of Quantum Dots in Living Mice SMALL Schipper, M. L., Iyer, G., Koh, A. L., Cheng, Z., Ebenstein, Y., Aharoni, A., Keren, S., Bentolila, L. A., Li, J., Rao, J., Chen, X., Banin, U., Wu, A. M., Sinclair, R., Weiss, S., Gambhir, S. S. 2009; 5 (1): 126-134

  Abstract

  This study evaluates the influence of particle size, PEGylation, and surface coating on the quantitative biodistribution of near-infrared-emitting quantum dots (QDs) in mice. Polymer- or peptide-coated 64Cu-labeled QDs 2 or 12 nm in diameter, with or without polyethylene glycol (PEG) of molecular weight 2000, are studied by serial micropositron emission tomography imaging and region-of-interest analysis, as well as transmission electron microscopy and inductively coupled plasma mass spectrometry. PEGylation and peptide coating slow QD uptake into the organs of the reticuloendothelial system (RES), liver and spleen, by a factor of 6-9 and 2-3, respectively. Small particles are in part renally excreted. Peptide-coated particles are cleared from liver faster than physical decay alone would suggest. Renal excretion of small QDs and slowing of RES clearance by PEGylation or peptide surface coating are encouraging steps toward the use of modified QDs for imaging living subjects.

  View details for DOI 10.1002/smll.200800003

  View details for Web of Science ID 000262895300019

  View details for PubMedID 19051182

  View details for PubMedCentralID PMC3084659

• Protein-Functionalized Synthetic Antiferromagnetic Nanoparticles for Biomolecule Detection and Magnetic Manipulation ANGEWANDTE CHEMIE-INTERNATIONAL EDITION Fu, A., Hu, W., Xu, L., Wilson, R. J., Yu, H., Osterfeld, S. J., Gambhir, S. S., Wang, S. X. 2009; 48 (9): 1620-1624

  Abstract

  Direct protein functionalization provides synthetic antiferromagnetic nanoparticles with high chemical specificity and multifunctionality. These nanoparticle-protein conjugates function as improved magnetic labels for biological detection experiments, and exhibit tunable responses to a small external magnetic field gradient, thus allowing the observation of distinctive single nanoparticle motion.

  View details for DOI 10.1002/anie.200803994

  View details for Web of Science ID 000263642300018

  View details for PubMedID 19156803

  View details for PubMedCentralID PMC2665302

• F-18-FDG PET/CT evaluation of patients with ovarian carcinoma NUCLEAR MEDICINE COMMUNICATIONS Iagaru, A. H., Mittra, E. S., McDougall, I. R., Quon, A., Gambhir, S. S. 2008; 29 (12): 1046-1051

  Abstract

  The role of F-FDG PET has been studied in ovarian carcinoma, but its sensitivity and specificity calculations are based on dedicated PET acquisition, not PET/CT in the majority of the published studies. Therefore, we were prompted to review our experience with PET/CT in the management of patients with ovarian carcinoma.This is a retrospective study of 43 women with ovarian carcinoma, 27-80 years old (average: 53.9+/-7.8), who had whole-body PET/CT at our institution from 1 January 2003 to 31 August 2006. We reviewed the patients' outcomes from medical records and compared them to the interpretation of the PET/CT scans. Sensitivity and specificity were calculated using a 2 x 2 table with pathology results (79.1% of the patients) or clinical follow-up (20.9% of the cases) as the 'gold standard'. Confidence interval (CI) estimations were performed using the Wilson score method.All patients had advanced stage ovarian cancer and the study was requested for re-staging. A total of 60 scans were performed: 30 patients had one scan, nine patients had two scans and four patients had three scans. The administered doses of F-FDG ranged from 381.1 to 769.6 MBq (average: 569.8+/-73.3). PET/CT had a sensitivity of 88.4% (95% CI: 75.1-95.4) and a specificity of 88.2% (95% CI: 64.4-97.9) for detection of ovarian cancer. The SUV max of the detected lesions ranged from 3 to 27 (average: 9.4+/-5.9). The CA-125 tumor marker ranged from 3 to 935 kU/ml (average: 265.2) in patients with positive scans and 4-139 kU/ml (average: 17.1) in patients with negative scans. This difference was statistically significant (P value: 0.0242).This study confirms the good results of F-FDG PET/CT for identification of residual/recurrent ovarian cancer, as well as for distant metastases localization. PET/CT should be an integral part in evaluation of patients with high-risk ovarian cancer or rising values of tumor markers (CA-125), prior to selection of the most appropriate therapy.

  View details for DOI 10.1097/MNM.0b013e32831089cb

  View details for Web of Science ID 000261164200004

  View details for PubMedID 18987524

  View details for PubMedCentralID PMC2651960

• Uptake kinetics and biodistribution of C-14-D-luciferin-a radiolabeled substrate for the firefly luciferase catalyzed bioluminescence reaction: impact on bioluminescence based reporter gene imaging EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Berger, F., Paulmurugan, R., Bhaumik, S., Gambhir, S. S. 2008; 35 (12): 2275-2285

  Abstract

  Firefly luciferase catalyzes the oxidative decarboxylation of D: -luciferin to oxyluciferin in the presence of cofactors, producing bioluminescence. This reaction is used in optical bioluminescence-based molecular imaging approaches to detect the expression of the firefly luciferase reporter gene. Biokinetics and distribution of the substrate most likely have a significant impact on levels of light signal and therefore need to be investigated.Benzene ring (14)C(U)-labeled D-luciferin was utilized. Cell uptake and efflux assays, murine biodistribution, autoradiography and CCD-camera based optical bioluminescence imaging were carried out to examine the in vitro and in vivo characteristics of the tracer in cell culture and in living mice respectively.Radiolabeled and unlabeled D-luciferin revealed comparable levels of light emission when incubated with equivalent amounts of the firefly luciferase enzyme. Cell uptake assays in pCMV-luciferase-transfected cells showed slow trapping of the tracer and relatively low uptake values (up to 22.9-fold higher in firefly luciferase gene-transfected vs. nontransfected cells, p = 0.0002). Biodistribution studies in living mice after tail-vein injection of (14)C-D-luciferin demonstrated inhomogeneous tracer distribution with early predominant high radioactivity levels in kidneys (10.6% injected dose [ID]/g) and liver (11.9% ID/g), followed at later time points by the bladder (up to 81.3% ID/g) and small intestine (6.5% ID/g), reflecting the elimination routes of the tracer. Kinetics and uptake levels profoundly differed when using alternate injection routes (intravenous versus intraperitoneal). No clear trapping of (14)C-D-luciferin in firefly luciferase-expressing tissues could be observed in vivo.The data obtained with (14)C-D-luciferin provide insights into the dynamics of D: -luciferin cell uptake, intracellular accumulation, and efflux. Results of the biodistribution and autoradiographic studies should be useful for optimizing and adapting optical imaging protocols to specific experimental settings when utilizing the firefly luciferase and D: -luciferin system.

  View details for DOI 10.1007/s00259-008-0870-6

  View details for Web of Science ID 000261654000015

  View details for PubMedID 18661130

• Ovarian Cancer Early Detection Claims Are Biased CLINICAL CANCER RESEARCH McIntosh, M., Anderson, G., Drescher, C., Hanash, S., Urban, N., Brown, P., Gambhir, S., Coukos, G., Laird, P. W., Nelson, B., Palmer, C. 2008; 14 (22): 7574

  View details for DOI 10.1158/1078-0432.CCR-08-0623

  View details for Web of Science ID 000261014500051

  View details for PubMedID 18948385

• Noninvasive Imaging of Therapeutic Gene Expression Using a Bidirectional Transcriptional Amplification Strategy MOLECULAR THERAPY Ray, S., Paulmurugan, R., Patel, M. R., Ahn, B. C., Wu, L., Carey, M., Gambhir, S. S. 2008; 16 (11): 1848-1856

  Abstract

  Promoters that limit transgene expression to tumors play a vital role in cancer gene therapy. Although tumor specific, the human Survivin promoter (pSurv) elicits low levels of transcription. A bidirectional two-step transcriptional amplification (TSTA) system was designed to enhance expression of the therapeutic gene (TG) tumor necrosis factor-alpha-related apoptosis-inducing ligand (TRAIL or TR) and the reporter gene firefly luciferase (FL) from pSurv. An adenoviral vector carrying the enhanced targeting apparatus (Ad-pSurv-TR-G8-FL) was tested for efficiency and specificity of gene expression in cells and in living animals. Compared to the one-step systems (Ad-pSurv-FL or Ad-pSurv-TR), the bidirectional TSTA system showed tenfold higher expression of both the therapeutic and the reporter gene and their expression correlated in cells (R(2) = 0.99) and in animals (R(2) = 0.67). Noninvasive quantitative monitoring of magnitude and time variation of TRAIL gene expression was feasible by bioluminescence imaging of the transcriptionally linked FL gene in xenograft tumors following intratumoral adenoviral injection. Moreover, the TSTA adenovirus maintained promoter specificity in nontarget tissues following tail vein administration. These studies demonstrate the potential of the bidirectional TSTA system to achieve high levels of gene expression from a weak promoter, while preserving specificity and the ability to image expression of the TG noninvasively.

  View details for DOI 10.1038/mt.2008.180

  View details for Web of Science ID 000260472600015

  View details for PubMedID 18766175

  View details for PubMedCentralID PMC3195556

• Adenoviral and Retroviral Transduction of the Thymidine Kinase PET Reporter Gene in Mesenchymal Stem Cells: Effect on Functional Parameters and Length of Expression Roelants, V., de Meester, C., Havaux, X., Tabilio, A., Gambhir, S. S., Di Ianni, M., Bertrand, L., Vanoverschelde, J. LIPPINCOTT WILLIAMS & WILKINS. 2008: S1039

  View details for Web of Science ID 000262104504134

• Transcriptional and Functional Profiling of Human Embryonic Stem Cell-Derived Cardiomyocytes PLOS ONE Cao, F., Wagner, R. A., Wilson, K. D., Xie, X., Fu, J., Drukker, M., Lee, A., Li, R. A., Gambhir, S. S., Weissman, I. L., Robbins, R. C., Wu, J. C. 2008; 3 (10)

  Abstract

  Human embryonic stem cells (hESCs) can serve as a potentially limitless source of cells that may enable regeneration of diseased tissue and organs. Here we investigate the use of human embryonic stem cell-derived cardiomyocytes (hESC-CMs) in promoting recovery from cardiac ischemia reperfusion injury in a mouse model. Using microarrays, we have described the hESC-CM transcriptome within the spectrum of changes that occur between undifferentiated hESCs and fetal heart cells. The hESC-CMs expressed cardiomyocyte genes at levels similar to those found in 20-week fetal heart cells, making this population a good source of potential replacement cells in vivo. Echocardiographic studies showed significant improvement in heart function by 8 weeks after transplantation. Finally, we demonstrate long-term engraftment of hESC-CMs by using molecular imaging to track cellular localization, survival, and proliferation in vivo. Taken together, global gene expression profiling of hESC differentiation enables a systems-based analysis of the biological processes, networks, and genes that drive hESC fate decisions, and studies such as this will serve as the foundation for future clinical applications of stem cell therapies.

  View details for DOI 10.1371/journal.pone.0003474

  View details for Web of Science ID 000265126100005

  View details for PubMedID 18941512

  View details for PubMedCentralID PMC2565131

• F-18 FDG PET/CT Evaluation of Osseous and Soft Tissue Sarcomas: Differences between Adult and Pediatric Patients Iagaru, A., Mittra, E., Quon, A., Jacobs, C., Marina, N., Gambhir, S. S. SPRINGER. 2008: S155

  View details for Web of Science ID 000208690700129

• Noninvasive Raman spectroscopy in living mice for evaluation of tumor targeting with carbon nanotubes NANO LETTERS Zavaleta, C., de la Zerda, A., Liu, Z., Keren, S., Cheng, Z., Schipper, M., Chen, X., Dai, H., Gambhir, S. S. 2008; 8 (9): 2800-2805

  Abstract

  An optimized noninvasive Raman microscope was used to evaluate tumor targeting and localization of single walled carbon nanotubes (SWNTs) in mice. Raman images were acquired in two groups of tumor-bearing mice. The control group received plain-SWNTs, whereas the experimental group received tumor targeting RGD-SWNTs intravenously. Raman imaging commenced over the next 72 h and revealed increased accumulation of RGD-SWNTs in tumor ( p < 0.05) as opposed to plain-SWNTs. These results support the development of a new preclinical Raman imager.

  View details for DOI 10.1021/nl801362a

  View details for Web of Science ID 000259140200034

  View details for PubMedID 18683988

  View details for PubMedCentralID PMC2910584

• Dual-targeted contrast agent for US assessment of tumor angiogenesis in vivo RADIOLOGY Willmann, J. K., Lutz, A. M., Paulmurugan, R., Patel, M. R., Chu, P., Rosenberg, J., Gambhir, S. S. 2008; 248 (3): 936-944

  Abstract

  To develop and validate a dual-targeted ultrasonographic (US) imaging agent with microbubbles (MBs) that attaches to both vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) and alpha(v)beta(3) integrin and to compare the US imaging signal obtained from dual-targeted MBs (MB(D)) with that from single-targeted MBs (MB(S)) in a murine model of tumor angiogenesis.Animal protocols were approved by the institutional Administrative Panel on Laboratory Animal Care. Single- and dual-targeted US imaging agents were prepared by attaching anti-VEGFR2, anti-alpha(v)beta(3) integrin, or both antibodies to the shell of perfluorocarbon-filled MBs. Binding specificities of targeted MBs compared with isotype-matched immunoglobulin G-labeled control MBs (MB(C)) and nontargeted nonlabeled MBs (MB(N)) were tested with VEGFR2-positive and alpha(v)beta(3) integrin-positive cells (mouse SVR cells) and control cells (mouse 4T1 cells). In vivo imaging signals of contrast material-enhanced US by using anti-VEGFR2-targeted MBs (MB(V)), anti-alpha(v)beta(3) integrin-targeted MBs (MB(I)), MB(D), and MB(C) were quantified in 49 mice bearing SK-OV-3 tumors (human ovarian cancer). Tumor tissue was stained for VEGFR2, alpha(v)beta(3) integrin, and CD31.Attachment of MB(D) to SVR cells (mean, 0.74 MBs per cell +/- 0.05 [standard deviation]) was significantly higher than attachment to 4T1 cells (mean, 0.04 +/- 0.03), and attachment to SVR cells was higher for MB(D) than for MB(V) (mean, 0.58 +/- 0.09), MB(I) (mean, 0.42 +/- 0.21), MB(C) (mean, 0.11 +/- 0.13), and MB(N) (mean, 0.01 +/- 0.01) (P < .05). Imaging signal in the murine tumor angiogenesis model was significantly higher (P < .001) for MB(D) (mean, 16.7 +/- 7.2) than for MB(V) (mean, 11.3 +/- 5.7), MB(I) (mean, 7.8 +/- 5.3), MB(C) (mean, 2.8 +/- 0.9), and MB(N) (mean, 1.1 +/- 0.4). Immunofluorescence confirmed expression of VEGFR2 and alpha(v)beta(3) integrin on tumor vasculature.Dual-targeted contrast-enhanced US directed at both VEGFR2 and alpha(v)beta(3) integrin improves in vivo visualization of tumor angiogenesis in a human ovarian cancer xenograft tumor model in mice.http://radiology.rsnajnls.org/cgi/content/full/248/3/936/DC1.

  View details for DOI 10.1148/radiol.2483072231

  View details for Web of Science ID 000258541500031

  View details for PubMedID 18710985

  View details for PubMedCentralID PMC2798094

• F-18-FDG-PET/CT evaluation of response to treatment in lymphoma: when is the optimal time for the first re-evaluation scan? HELLENIC JOURNAL OF NUCLEAR MEDICINE Iagaru, A., Wang, Y., Mari, C., Quon, A., Goris, M. L., Horning, S., Gambhir, S. S. 2008; 11 (3): 153-156

  Abstract

  Assessing the response to treatment as soon after treatment initiation is one of the key reasons for imaging lymphoma patients. The optimal time after initiating treatment for assessing response to treatment has yet to be determined. Therefore, we were prompted to review our experience with serial (18)F-FDG PET/CT in patients undergoing treatment for Hodgkin's disease (HD) and non Hodgkin's lymphoma (NHL). This is a retrospective study (Feb 2003 - Oct 2004) of 20 patients, 11 men and 9 women, with age range of 7-75 years with diagnosis of HD (10) and NHL (10), who had PET/CT at our institution prior, during and at the completion of therapy. Restaging PET/CT was done after 2 cycles of chemotherapy in 10 patients (group A) and after 4 cycles of chemotherapy in 10 pts (group B). A total of 60 scans were reviewed. The DeltaSUV from baseline to first PET/CT was on average 67.6% in group A and 75.1% in group B. This had no statistical significance (P value: 0.31). The DeltaSUV from baseline to post-therapy PET/CT was on average 72.9% in group A and 79.8% in group B. This difference also had no statistical significance (P value: 0.24). The correlation coefficient was 0.98 in group A and 0.80 in group B. Results of PET/CT after 2 cycles of chemotherapy did not statistically differ from the results of PET/CT after 4 cycles of chemotherapy. These results need to be confirmed in larger, prospective, randomized trials.

  View details for Web of Science ID 000262093600003

  View details for PubMedID 19081857

• Carbon nanotubes as photoacoustic molecular imaging agents in living mice NATURE NANOTECHNOLOGY de la Zerda, A., Zavaleta, C., Keren, S., Vaithilingam, S., Bodapati, S., Liu, Z., Levi, J., Smith, B. R., Ma, T., Oralkan, O., Cheng, Z., Chen, X., Dai, H., Khuri-Yakub, B. T., Gambhir, S. S. 2008; 3 (9): 557-562

  Abstract

  Photoacoustic imaging of living subjects offers higher spatial resolution and allows deeper tissues to be imaged compared with most optical imaging techniques. As many diseases do not exhibit a natural photoacoustic contrast, especially in their early stages, it is necessary to administer a photoacoustic contrast agent. A number of contrast agents for photoacoustic imaging have been suggested previously, but most were not shown to target a diseased site in living subjects. Here we show that single-walled carbon nanotubes conjugated with cyclic Arg-Gly-Asp (RGD) peptides can be used as a contrast agent for photoacoustic imaging of tumours. Intravenous administration of these targeted nanotubes to mice bearing tumours showed eight times greater photoacoustic signal in the tumour than mice injected with non-targeted nanotubes. These results were verified ex vivo using Raman microscopy. Photoacoustic imaging of targeted single-walled carbon nanotubes may contribute to non-invasive cancer imaging and monitoring of nanotherapeutics in living subjects.

  View details for DOI 10.1038/nnano.2008.231

  View details for Web of Science ID 000259013100014

  View details for PubMedID 18772918

  View details for PubMedCentralID PMC2562547

• BIOT 45-Cystine knot polypeptides engineered for high affinity integrin binding: A new class of in vivo molecular imaging agents Kimura, R. H., Levin, A. M., Cheng, Z., Gambhir, S., Cochran, J. R. AMER CHEMICAL SOC. 2008

  View details for Web of Science ID 000270256302136

• Cancer screening: A mathematical model relating secreted blood biomarker levels to tumor sizes PLOS MEDICINE Lutz, A. M., Willmann, J. K., Cochran, F. V., Ray, P., Gambhir, S. S. 2008; 5 (8): 1287-1297

  Abstract

  Increasing efforts and financial resources are being invested in early cancer detection research. Blood assays detecting tumor biomarkers promise noninvasive and financially reasonable screening for early cancer with high potential of positive impact on patients' survival and quality of life. For novel tumor biomarkers, the actual tumor detection limits are usually unknown and there have been no studies exploring the tumor burden detection limits of blood tumor biomarkers using mathematical models. Therefore, the purpose of this study was to develop a mathematical model relating blood biomarker levels to tumor burden.Using a linear one-compartment model, the steady state between tumor biomarker secretion into and removal out of the intravascular space was calculated. Two conditions were assumed: (1) the compartment (plasma) is well-mixed and kinetically homogenous; (2) the tumor biomarker consists of a protein that is secreted by tumor cells into the extracellular fluid compartment, and a certain percentage of the secreted protein enters the intravascular space at a continuous rate. The model was applied to two pathophysiologic conditions: tumor biomarker is secreted (1) exclusively by the tumor cells or (2) by both tumor cells and healthy normal cells. To test the model, a sensitivity analysis was performed assuming variable conditions of the model parameters. The model parameters were primed on the basis of literature data for two established and well-studied tumor biomarkers (CA125 and prostate-specific antigen [PSA]). Assuming biomarker secretion by tumor cells only and 10% of the secreted tumor biomarker reaching the plasma, the calculated minimally detectable tumor sizes ranged between 0.11 mm(3) and 3,610.14 mm(3) for CA125 and between 0.21 mm(3) and 131.51 mm(3) for PSA. When biomarker secretion by healthy cells and tumor cells was assumed, the calculated tumor sizes leading to positive test results ranged between 116.7 mm(3) and 1.52 x 10(6) mm(3) for CA125 and between 27 mm(3) and 3.45 x 10(5) mm(3) for PSA. One of the limitations of the study is the absence of quantitative data available in the literature on the secreted tumor biomarker amount per cancer cell in intact whole body animal tumor models or in cancer patients. Additionally, the fraction of secreted tumor biomarkers actually reaching the plasma is unknown. Therefore, we used data from published cell culture experiments to estimate tumor cell biomarker secretion rates and assumed a wide range of secretion rates to account for their potential changes due to field effects of the tumor environment.This study introduced a linear one-compartment mathematical model that allows estimation of minimal detectable tumor sizes based on blood tumor biomarker assays. Assuming physiological data on CA125 and PSA from the literature, the model predicted detection limits of tumors that were in qualitative agreement with the actual clinical performance of both biomarkers. The model may be helpful in future estimation of minimal detectable tumor sizes for novel proteomic biomarker assays if sufficient physiologic data for the biomarker are available. The model may address the potential and limitations of tumor biomarkers, help prioritize biomarkers, and guide investments into early cancer detection research efforts.

  View details for DOI 10.1371/journal.pmed.0050170

  View details for Web of Science ID 000258739200018

  View details for PubMedID 18715113

  View details for PubMedCentralID PMC2517618

• A human estrogen receptor (ER)alpha mutation with differential responsiveness to nonsteroidal ligands: Novel approaches for studying mechanism of ER action MOLECULAR ENDOCRINOLOGY Paulmurugan, R., Tamrazi, A., Katzenellenbogen, J. A., Katzenellenbogen, B. S., Gambhir, S. S. 2008; 22 (7): 1552-1564

  Abstract

  Estrogens, acting through the estrogen receptors (ERs), play crucial roles in regulating the function of reproductive and other systems under physiological and pathological conditions. ER activity in regulating target genes is modulated by the binding of both steroidal and synthetic nonsteroidal ligands, with ligand binding inducing ERs to adopt various conformations that control their interactions with transcriptional coregulators. Previously, we developed an intramolecular folding sensor with a mutant form of ERalpha (ER(G521T)) that proved to be essentially unresponsive to the endogenous ligand 17beta-estradiol, yet responded very well to certain synthetic ligands. In this study, we have characterized this G521T-ER mutation in terms of the potency and efficacy of receptor response toward several steroidal and nonsteroidal ligands in two different ways: directly, by ligand effects on mutant ER conformation (by the split-luciferase complementation system), and indirectly, by ligand effects on mutant ER transactivation. Full-length G521T-ER shows no affinity for estradiol and does not activate an estrogen-responsive reporter gene. The synthetic pyrazole agonist ligand propyl-pyrazole-triol is approximately 100-fold more potent than estradiol in inducing intramolecular folding and reporter gene transactivation with the mutant ER, whereas both ligands have high potency on wild-type ER. This estradiol-unresponsive mutant ER can also specifically highlight the agonistic property of the selective ER modulator, 4-hydroxytamoxifen, by reporter gene transactivation, even in the presence of estradiol, and it can exert a dominant-negative effect on estrogen-stimulated wild-type ER. This system provides a model for ER-mutants that show differential ligand responsiveness to gene activation to gain insight into the phenomenon of hormone resistance observed in endocrine therapies of ER-positive breast cancers.

  View details for DOI 10.1210/me.2007-0570

  View details for PubMedID 18451095

• Direct site-specific radiolabeling of an affibody protein with 4-[F-18]fluorobenzaldehyde via oxime chemistry MOLECULAR IMAGING AND BIOLOGY Namavari, M., De Jesus, O. P., Cheng, Z., De, A., Kovacs, E., Levi, J., Zhang, R., Hoerner, J. K., Grade, H., Syud, F. A., Gambhir, S. S. 2008; 10 (4): 177-181

  Abstract

  In this study, we introduce a methodology for preparing 18F-labeled Affibody protein, specifically 18F-Anti-HER2 dimeric Affibody (14 kDa), for in vivo imaging of HER2neu with positron emission tomography (PET).We have used 4-[18F]fluorobenzaldehyde as a synthon to prepare 18F-Anti-HER2 Affibody. Aminooxy-functionalized Affibody (Anti-HER2-ONH2) was incubated with 4-[18F]fluorobenzaldehyde in ammonium acetate buffer at pH 4 in the presence of methanol at 70 degrees C for 15 min. The resulting 18F-labeled Affibody molecule was evaluated as a PET probe in xenograft models expressing HER2.We have successfully prepared 18F-Anti-HER2 dimeric Affibody (14 kDa), N-(4-[18F]fluorobenzylidine)oxime-Anti-HER2 Affibody, [18F]FBO-Anti-HER2, in 26-30% radiochemical yields (decay corrected). High-contrast small-animal PET images with relatively moderate tumor uptake (1.79 +/- 0.40% ID/g) were observed for the 18F-Anti-HER2 Affibody.Site-specific 18F-labeled Affibody against HER2 has been synthesized via chemoselective oxime formation between an aminooxy-functionalized Affibody and 18F-fluorobenzaldehyde. The results have implications for radiolabeling of other affibodies and macromolecules and should also be important for advancing Affibody imaging with PET.

  View details for DOI 10.1007/s11307-008-0142-7

  View details for Web of Science ID 000256929100001

• Direct site-specific radiolabeling of an Affibody protein with 4-[18F]fluorobenzaldehyde via oxime chemistry. Molecular imaging and biology Namavari, M., Padilla De Jesus, O., Cheng, Z., De, A., Kovacs, E., Levi, J., Zhang, R., Hoerner, J. K., Grade, H., Syud, F. A., Gambhir, S. S. 2008; 10 (4): 177-181

  Abstract

  In this study, we introduce a methodology for preparing 18F-labeled Affibody protein, specifically 18F-Anti-HER2 dimeric Affibody (14 kDa), for in vivo imaging of HER2neu with positron emission tomography (PET).We have used 4-[18F]fluorobenzaldehyde as a synthon to prepare 18F-Anti-HER2 Affibody. Aminooxy-functionalized Affibody (Anti-HER2-ONH2) was incubated with 4-[18F]fluorobenzaldehyde in ammonium acetate buffer at pH 4 in the presence of methanol at 70 degrees C for 15 min. The resulting 18F-labeled Affibody molecule was evaluated as a PET probe in xenograft models expressing HER2.We have successfully prepared 18F-Anti-HER2 dimeric Affibody (14 kDa), N-(4-[18F]fluorobenzylidine)oxime-Anti-HER2 Affibody, [18F]FBO-Anti-HER2, in 26-30% radiochemical yields (decay corrected). High-contrast small-animal PET images with relatively moderate tumor uptake (1.79 +/- 0.40% ID/g) were observed for the 18F-Anti-HER2 Affibody.Site-specific 18F-labeled Affibody against HER2 has been synthesized via chemoselective oxime formation between an aminooxy-functionalized Affibody and 18F-fluorobenzaldehyde. The results have implications for radiolabeling of other affibodies and macromolecules and should also be important for advancing Affibody imaging with PET.

  View details for DOI 10.1007/s11307-008-0142-7

  View details for PubMedID 18481153

• A generalizable strategy for imaging pre-mRNA levels in living subjects using spliceosome-mediated RNA trans-splicing JOURNAL OF NUCLEAR MEDICINE Walls, Z. F., Puttaraju, M., Temple, G. F., Gambhir, S. S. 2008; 49 (7): 1146-1154

  Abstract

  Molecular imaging of gene expression is currently hindered by the lack of a generalizable platform for probe design. For any gene of interest, a probe that targets protein levels must often be generated empirically. Targeting gene expression at the level of mRNA, however, would allow probes to be built on the basis of sequence information alone. Presented here is a class of generalizable probes that can image pre-mRNA in a sequence-specific manner, using signal amplification and a facile method of delivery.Pre-trans-splicing molecules (PTMs) were engineered to capitalize on the phenomenon of spliceosome-mediated RNA trans-splicing. Using a modular binding domain that confers specificity by base-pair complementarity to the target pre-mRNA, PTMs were designed to target a chimeric target mini gene and trans-splice the Renilla luciferase gene onto the end of the target. PTMs and target genes were transfected in cell culture and assessed by luciferase assay, reverse-transcriptase polymerase chain reaction, Western blot, and rapid analysis of 5' cDNA ends. PTMs and target genes were also assessed in vivo by hydrodynamic delivery in mice.Efficiency and specificity of the trans-splicing reaction were found to vary depending on the binding domain length and structure. Specific trans-splicing was observed in living animals (P = 0.0862, Kruskal-Wallis test).Described here is a model system used to demonstrate the feasibility of spliceosome-mediated RNA trans-splicing for imaging gene expression at the level of pre-mRNA using optical imaging techniques in living animals. The experiments reported here show proof of principle for a generalizable imaging probe against RNA that can amplify signal on detection and be delivered using existing gene delivery methodology.

  View details for DOI 10.2967/jnumed.107.047662

  View details for Web of Science ID 000257599700021

  View details for PubMedID 18552150

• Molecular imaging of cancer: From molecules to humans - Introduction JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 2008; 49: 1S-4S

  View details for DOI 10.2967/jnumed.108.053751

  View details for Web of Science ID 000256491100001

  View details for PubMedID 18523062

• Molecular Imaging of Hypoxia-Inducible Factor 1 alpha and von Hippel-Lindau Interaction in Mice MOLECULAR IMAGING Choi, C. Y., Chau, D. A., Paulmurugan, R., Sutphin, P. D., Le, Q., Koong, A. C., Zundel, W., Gambhir, S. S., Giaccia, A. J. 2008; 7 (3): 139-146

  Abstract

  Tumor hypoxia plays a crucial role in tumorigenesis. Under hypoxia, hypoxia-inducible factor 1 alpha (HIF-1 alpha) regulates activation of genes promoting malignant progression. Under normoxia, HIF-1 alpha is hydroxylated on prolines 402 and 564 and is targeted for ubiquitin-mediated degradation by interacting with the von Hippel-Lindau protein complex (pVHL). We have developed a novel method of studying the interaction between HIF-1 alpha and pVHL using the split firefly luciferase complementation-based bioluminescence system in which HIF-1 alpha and pVHL are fused to amino-terminal and carboxy-terminal fragments of the luciferase, respectively. We demonstrate that hydroxylation-dependent interaction between the HIF-1 alpha and pVHL leads to complementation of the two luciferase fragments, resulting in bioluminescence in vitro and in vivo. Complementation-based bioluminescence is diminished when mutant pVHLs with decreased affinity for binding HIF-1 alpha are used. This method represents a new approach for studying interaction of proteins involved in the regulation of protein degradation.

  View details for DOI 10.2310/7290.2008.00017

  View details for Web of Science ID 000260954700004

  View details for PubMedID 19123984

• Cell-free metabolic engineering promotes high-level production of bioactive Gaussia princeps luciferase METABOLIC ENGINEERING Goerke, A. R., Loening, A. M., Gambhir, S. S., Swartz, J. R. 2008; 10 (3-4): 187-200

  Abstract

  Due to its small size and intense luminescent signal, Gaussia princeps luciferase (GLuc) is attractive as a potential imaging agent in both cell culture and small animal research models. However, recombinant GLuc production using in vivo techniques has only produced small quantities of active luciferase, likely due to five disulfide bonds being required for full activity. Cell-free biology provides the freedom to control both the catalyst and chemical compositions in biological reactions, and we capitalized on this to produce large amounts of highly active GLuc in cell-free reactions. Active yields were improved by mutating the cell extract source strain to reduce proteolysis, adjusting reaction conditions to enhance oxidative protein folding, further activating energy metabolism, and encouraging post-translational activation. This cell-free protein synthesis procedure produced 412mug/mL of purified GLuc, relative to 5mug/mL isolated for intracellular Escherichia coli expression. The cell-free product had a specific activity of 4.2x10(24)photons/s/mol, the highest reported activity for any characterized luciferase.

  View details for DOI 10.1016/j.ymben.2008.04.001

  View details for PubMedID 18555198

• Imaging of VEGF receptor in a rat myocardial infarction model using PET JOURNAL OF NUCLEAR MEDICINE Rodriguez-Porcel, M., Cai, W., Gheysens, O., Willmann, J. K., Chen, K., Wang, H., Chen, I. Y., He, L., Wu, J. C., Li, Z., Mohamedali, K. A., Kim, S., Rosenblum, M. G., Chen, X., Gambhir, S. S. 2008; 49 (4): 667-673

  Abstract

  Myocardial infarction (MI) leads to left ventricular (LV) remodeling, which leads to the activation of growth factors such as vascular endothelial growth factor (VEGF). However, the kinetics of a growth factor's receptor expression, such as VEGF, in the living subject has not yet been described. We have developed a PET tracer (64Cu-DOTA-VEGF121 [DOTA is 1,4,7,10-tetraazadodecane-N,N',N'',N'''-tetraacetic acid]) to image VEGF receptor (VEGFR) expression after MI in the living subject.In Sprague-Dawley rats, MI was induced by ligation of the left coronary artery and confirmed by ultrasound (n = 8). To image and study the kinetics of VEGFRs, 64Cu-DOTA-VEGF121 PET scans were performed before MI induction (baseline) and on days 3, 10, 17, and 24 after MI. Sham-operated animals served as controls (n = 3).Myocardial origin of the 64Cu-DOTA-VEGF121 signal was confirmed by CT coregistration and autoradiography. VEGFR specificity of the 64Cu-DOTA-VEGF121 probe was confirmed by in vivo use of a 64Cu-DOTA-VEGFmutant. Baseline myocardial uptake of 64Cu-DOTA-VEGF121 was minimal (0.30 +/- 0.07 %ID/g [percentage injected dose per gram of tissue]); it increased significantly after MI (day 3, 0.97 +/- 0.05 %ID/g; P < 0.05 vs. baseline) and remained elevated for 2 wk (up to day 17 after MI), after which time it returned to baseline levels.We demonstrate the feasibility of imaging VEGFRs in the myocardium. In summary, we imaged and described the kinetics of 64Cu-DOTA-VEGF121 uptake in a rat model of MI. Studies such as the one presented here will likely play a major role when studying pathophysiology and assessing therapies in different animal models of disease and, potentially, in patients.

  View details for DOI 10.2967/jnumed.107.040576

  View details for PubMedID 18375924

• Configurations of a two-tiered amplified gene expression system in adenoviral vectors designed to improve the specificity of in vivo prostate cancer imaging GENE THERAPY Sato, M., Figueiredo, M. L., Burton, J. B., Johnson, M., Chen, M., Powell, R., Gambhir, S. S., Carey, M., Wu, L. 2008; 15 (8): 583-593

  Abstract

  Effective treatment for recurrent, disseminated prostate cancer is notably limited. We have developed adenoviral vectors with a prostate-specific two-step transcriptional amplification (TSTA) system that would express therapeutic genes at a robust level to target metastatic disease. The TSTA system employs the prostate-specific antigen (PSA) promoter/enhancer to drive a potent synthetic activator, which in turn activates the expression of the therapeutic gene. In this study, we explored different configurations of this bipartite system and discovered that physical separation of the two TSTA components into E1 and E3 regions of adenovirus was able to enhance androgen regulation and cell-discriminatory expression. The TSTA vectors that express imaging reporter genes were assessed by noninvasive imaging technologies in animal models. The improved selectivity of the E1E3 configured vector was reflected in silenced ectopic expression in the lung. Significantly, the enhanced specificity of the E1E3 vector enabled the detection of lung metastasis of prostate cancer. An E1E3 TSTA vector that expresses the herpes simplex virus thymidine kinase gene can effectively direct positron emission tomography (PET) imaging of the tumor. The prostate-targeted gene delivery vectors with robust and cell-specific expression capability will advance the development of safe and effective imaging guided therapy for recurrent metastatic stages of prostate cancer.

  View details for DOI 10.1038/gt.2008.19

  View details for Web of Science ID 000254561200004

  View details for PubMedID 18305574

  View details for PubMedCentralID PMC2798063

• Perspectives of molecular imaging and radioimmunotherapy in lymphoma RADIOLOGIC CLINICS OF NORTH AMERICA Iagaru, A., Goris, M. L., Gambhir, S. S. 2008; 46 (2): 243-252

  Abstract

  Successful treatment of Hodgkin lymphomas and non-Hodgkin lymphomas depends on accurate staging and prognostic estimations, as well as evaluation of response to therapy as early after initiation as possible. We focus on several aspects of molecular imaging and therapy that affect the management of patients who have lymphoma. First, we review prior use of gallium-67 citrate for evaluation of lymphoma patients, mainly from a historical perspective, since it was the mainstream lymphoma functional imaging tracer for decades. Next, we review current clinical uses of 18F Fluoro-2-Deoxyglucose (18F FDG) PET and PET/CT for evaluation of lymphoma patients and use of radioimmunotherapy in lymphoma. Finally, we discuss advances in molecular imaging that may herald the next generation of PET radiotracers after 18F FDG.

  View details for DOI 10.1016/j.rcl.2008.03.007

  View details for Web of Science ID 000258543500006

  View details for PubMedID 18619379

• Noninvasive molecular neuroimaging using reporter genes: Part II, experimental, current, and future applications AMERICAN JOURNAL OF NEURORADIOLOGY Massoud, T. F., Singh, A., Gambhir, S. S. 2008; 29 (3): 409-418

  Abstract

  In this second article, we review the various strategies and applications that make use of reporter genes for molecular imaging of the brain in living subjects. These approaches are emerging as valuable tools for monitoring gene expression in diverse applications in laboratory animals, including the study of gene-targeted and trafficking cells, gene therapies, transgenic animals, and more complex molecular interactions within the central nervous system. Further development of more sensitive and selective reporters, combined with improvements in detection technology, will consolidate the position of in vivo reporter gene imaging as a versatile technique for greater understanding of intracellular biologic processes and underlying molecular neuropathology and will potentially establish a future role in the clinical management of patients with neurologic diseases.

  View details for DOI 10.3174/ajnr.A0863

  View details for Web of Science ID 000254066700002

  View details for PubMedID 18272565

• Initial evaluation of F-18-fluorothymidine (FLT) PET/CT scanning for primary pancreatic cancer EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Quon, A., Chang, S. T., Chin, F., Kamaya, A., Dick, D. W., Loo, B. W., Gambhir, S. S., Koong, A. C. 2008; 35 (3): 527-531

  Abstract

  The aim of this study was to evaluate the potential of (18)F-fluorothymidine (FLT) PET/CT for imaging pancreatic adenocarcinoma.This was a pilot study of five patients (four males, one female) with newly diagnosed and previously untreated pancreatic adenocarcinoma. Patients underwent FLT PET/CT, (18)F-fluorodeoxyglucose (FDG) PET/CT, and contrast-enhanced CT scanning before treatment. The presence of cancer was confirmed by histopathological analysis at the time of scanning in all five patients. The degree of FLT and FDG uptake at the primary tumor site was assessed using visual interpretation and semi-quantitative SUV analyses.The primary tumor size ranged from 2.5 x 2.8 cm to 3.5 x 7.0 cm. The SUV of FLT uptake within the primary tumor ranged from 2.1 to 3.1. Using visual interpretation, the primary cancer could be detected from background activity in two of five patients (40%) on FLT PET/CT. By comparison, FDG uptake was higher in each patient with a SUV range of 3.4 to 10.8, and the primary cancer could be detected from background in all five patients (100%).In this pilot study of five patients with primary pancreatic adenocarcinoma, FLT PET/CT scanning showed poor lesion detectability and relatively low levels of radiotracer uptake in the primary tumor.

  View details for DOI 10.1007/s00259-007-0630-z

  View details for Web of Science ID 000254402800010

  View details for PubMedID 17960376

• Monitoring of the biological response to murine Hindlimb ischemia with Cu-64-labeled vascular endothelial growth factor-121 positron emission tomography CIRCULATION Willmann, J. K., Chen, K., Wang, H., Paulmurugan, R., Rollins, M., Cai, W., Wang, D. S., Chen, I. Y., Gheysens, O., Rodriguez-Porcel, M., Chen, X., Gambhir, S. S. 2008; 117 (7): 915-922

  Abstract

  Vascular endothelial growth factor-121 (VEGF121), an angiogenic protein secreted in response to hypoxic stress, binds to VEGF receptors (VEGFRs) overexpressed on vessels of ischemic tissue. The purpose of this study was to evaluate 64Cu-VEGF121 positron emission tomography for noninvasive spatial, temporal, and quantitative monitoring of VEGFR2 expression in a murine model of hindlimb ischemia with and without treadmill exercise training.64Cu-labeled VEGF121 and a VEGF mutant were tested for VEGFR2 binding specificity in cell culture. Mice (n=58) underwent unilateral ligation of the femoral artery, and postoperative tissue ischemia was assessed with laser Doppler imaging. Longitudinal VEGFR2 expression in exercised and nonexercised mice was quantified with 64Cu-VEGF121 positron emission tomography at postoperative day 8, 15, 22, and 29 and correlated with postmortem gamma-counting. Hindlimbs were excised for immunohistochemistry, Western blotting, and microvessel density measurements. Compared with the VEGF mutant, VEGF121 showed specific binding to VEGFR2. Perfusion in ischemic hindlimbs fell to 9% of contralateral hindlimb on postoperative day 1 and recovered to 82% on day 29. 64Cu-VEGF121 uptake in ischemic hindlimbs increased significantly (P < 0.001) from a control level of 0.61+/-0.17% ID/g (percentage of injected dose per gram) to 1.62+/-0.35% ID/g at postoperative day 8, gradually decreased over the following 3 weeks (0.59+/-0.14% ID/g at day 29), and correlated with gamma-counting (R2 = 0.99). Compared with nonexercised mice, 64Cu-VEGF121 uptake was increased significantly (P < or = 0.0001) in exercised mice (at day 15, 22, and 29) and correlated with VEGFR2 levels as obtained by Western blotting (R2 = 0.76). Ischemic hindlimb tissue stained positively for VEGFR2. In exercised mice, microvessel density was increased significantly (P<0.001) compared with nonexercised mice.64Cu-VEGF121 positron emission tomography allows longitudinal spatial and quantitative monitoring of VEGFR2 expression in murine hindlimb ischemia and indirectly visualizes enhanced angiogenesis stimulated by treadmill exercise training.

  View details for DOI 10.1161/CIRCULATIONAHA.107.733220

  View details for PubMedID 18250264

• Reporter gene imaging following percutaneous delivery in swine - Moving toward clinical applications JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY Rodriguez-Porcel, M., Brinton, T. J., Chen, I. Y., Gheysens, O., Lyons, J., Ikeno, F., Willmann, J. K., Wu, L., Wu, J. C., Yeung, A. C., Yock, P., Gambhir, S. S. 2008; 51 (5): 595-597

  View details for DOI 10.1016/j.jacc.2007.08.063

  View details for Web of Science ID 000252908600013

  View details for PubMedID 18237691

  View details for PubMedCentralID PMC2853907

• Noninvasive molecular neuroimaging using reporter genes: Part I, principles revisited AMERICAN JOURNAL OF NEURORADIOLOGY Massoud, T. F., Singh, A., Gambhir, S. S. 2008; 29 (2): 229-234

  Abstract

  In this first article, we review the basic principles of using reporter genes for molecular imaging of the brain in living subjects. This approach is emerging as a valuable tool for monitoring gene expression in diverse applications in laboratory animals, including the study of gene-targeted and trafficking cells, gene therapies, transgenic animals, and more complex molecular interactions within the central nervous system. Further development of more sensitive and selective reporters, combined with improvements in detection technology, will consolidate the position of in vivo reporter gene imaging as a versatile method for greater understanding of intracellular biologic processes and underlying molecular neuropathology and will potentially establish a future role in the clinical management of patients with neurologic diseases.

  View details for DOI 10.3174/ajnr.A0864

  View details for Web of Science ID 000253345200011

  View details for PubMedID 18024575

• Development of a bicistronic vector for multimodality imaging of estrogen receptor activity in a breast cancer model: preliminary application EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Ottobrini, L., Ciana, P., Moresco, R., Lecchi, M., Belloli, S., Martelli, C., Todde, S., Fazio, F., Gambhir, S., Maggi, A., Lucignani, G. 2008; 35 (2): 365–78

  Abstract

  The aim of this study was to develop a cellular model for the concurrent imaging of reporter genes expression by using positron emission tomography (PET) and bioluminescence imaging (BLI) for the assessment of estrogen receptor activity in vivo in a breast cancer model.Two reporters were chosen: a mutated form of the dopaminergic D2 receptor (D(2)R80A) for PET imaging, and the Firefly Luciferase for BLI. The presence of an IRES sequence between the two reporters ensured the coordinated expression driven by the same regulatory sequence containing an estrogen responsive element (ERE). To prevent chromatin effects on reporter expression, the construct was flanked by insulator sequences (Matrix Attachment Region, MAR).In vitro studies showed that the vector was efficient in coordinating the expression of the two genes. Moreover, stably transfected cells implanted in recipient animals maintained their capacity to express the reporters and react to systemic treatments permitting the in vivo study of ERs activity by PET and BLI imaging. In vitro expression analysis after long-term treatments showed different behaviour of the two reporter proteins in monitoring estrogen-dependent transcription outlining the importance of multi-reporter systems. With this model, PET and BLI can be applied to the concurrent evaluation of gene expression induced by estrogen and its analogues by using a bicistronic construct.The combined features of rapid, sensitive, sequential BLI and tomographic and quantitative PET imaging will allow the use of this strategy for the in vivo evaluation of molecular processes also for pharmacodynamic studies.

  View details for DOI 10.1007/s00259-007-0578-z

  View details for Web of Science ID 000252474400018

  View details for PubMedID 17926035

• US imaging of tumor angiogenesis with microbubbles targeted to vascular endothelial growth factor receptor type 2 in mice RADIOLOGY Willmann, J. K., Paulmurugan, R., Chen, K., Gheysens, O., Rodriguez-Porcel, M., Lutz, A. M., Chen, I. Y., Chen, X., Gambhir, S. S. 2008; 246 (2): 508-518

  Abstract

  To prospectively evaluate contrast material-enhanced ultrasonography (US) with microbubbles targeted to vascular endothelial growth factor receptor type 2 (VEGFR2) for imaging tumor angiogenesis in two murine tumor models.Animal protocols were approved by the Institutional Administrative Panel on Laboratory Animal Care. A US contrast agent, consisting of encapsulated gaseous microbubbles, was developed specifically to bind to VEGFR2 (by using anti-VEGFR2 antibodies and biotin-streptavidin interaction) which is up-regulated on endothelial cells of tumor blood vessels. VEGFR2-targeted microbubbles (MB(V)), control microbubbles (MB(C)), and nonlabeled microbubbles (MB(N)) were tested for binding specificity on cells expressing VEGFR2 (mouse angiosarcoma SVR cells) and control cells (mouse skeletal myoblast C2C12 cells). Expression of mouse VEGFR2 in culture cells was tested with immunocytochemical and Western blot analysis. Contrast-enhanced US imaging with MB(V) and MB(C) was performed in 28 tumor-bearing nude mice (mouse angiosarcoma, n = 18; rat malignant glioma, n = 10). Differences were calculated by using analysis of variance.In cell culture, adherence of MB(V) on SVR cells (2.1 microbubbles per SVR cell) was significantly higher than adherence of control microbubbles (0.01-0.10 microbubble per SVR cell; P < .001) and significantly more MB(V) attached to SVR cells than to C2C12 cells (0.15 microbubble per C2C12 cell; P < .001). In vivo, contrast-enhanced US imaging showed significantly higher average video intensity when using MB(V) compared with MB(C) for angiosarcoma and malignant glioma tumors (P < .001). Results of immunohistochemical analysis confirmed VEGFR2 expression on vascular endothelial cells of both tumor types.US imaging with contrast microbubbles targeted to VEGFR2 allows noninvasive visualization of VEGFR2 expression in tumor vessels in mice.

  View details for DOI 10.1148/radio1.2462070536

  View details for Web of Science ID 000252796300021

  View details for PubMedID 18180339

• 90Y Ibritumomal therapy in refractory non-Hodgkins lymphoma: Observations from 111 in-ibritumomab pre-treatment imaging Journal of Nuclear Medicine Iagaru A, Gambhir SS, Goris M 2008; 49 (11): 1809-1812
• Molecular Imaging of PET Reporter Gene Expression Molecular Imaging II Min, J., Gambhir, S. S. edited by Semmler, W., Schwaiger, M. Springer Verlag . 2008: 277–303
• Applications of Lentiviral Vectors in Non-Invasive Molecular Imaging Gene Therapy Protocols De, A., Yaghoubi, S., Gambhir, S. S. 2008; 3: 177–202
• Perspectives of Molecular Imaging and Radio Immunotherapy in Lymphoma Radiologic Clinics of North America Iagaru, A., Goris, M., Gambhir, S. S. 2008: 243–252
• Molecular Imaging of Cancer: From Molecules to Humans. Introduction. Molecular Imaging of Cancer: From Molecules to Humans. Introduction. Gambhir, S. 2008; 49: 1S-4S
• Use of Bioluminescent Imaging to Assay the Transplantation of Immortalized Human Fetal Hepatocytes Into Mice CELL TRANSPLANTATION Choi, M. S., Catana, A. M., Wu, J., Kim, Y. S., Yoon, S. J., Borowsky, A. D., Gambhir, S. S., Gupta, S., Zern, M. A. 2008; 17 (8): 899-909

  Abstract

  Noninvasive serial monitoring of the fate of transplanted cells would be invaluable to evaluate the potential therapeutic use of human hepatocyte transplantation. Therefore, we assessed the feasibility of bioluminescent imaging using double or triple fusion lentiviral vectors in a NOD-SCID mouse model transplanted with immortalized human fetal hepatocytes. Lentiviral vectors driven by the CMV promoter were constructed carrying reporter genes: firefly luciferase and green fluorescence protein with or without herpes simplex virus type 1 thymidine kinase. Human fetal hepatocytes immortalized by telomerase reconstitution (FH-hTERT) were successfully transduced with either of these fusion vectors. Two million stably transduced cells selected by fluorescence-activated cell sorting were injected into the spleens of NOD-SCID mice pretreated with methylcholanthrene and monocrotaline. The transplanted mice were serially imaged with a bioluminescence charged-coupled device camera after D-luciferin injection. Bioluminescence signal intensity was highest on day 3 (6.10 +/- 2.02 x 10(5) p/s/cm2/sr, mean +/- SEM), but decreased to 2.26 +/- 1.54 x 10(5) and 7.47 +/- 3.09 x 10(4) p/s/cm2/sr on day 7 and 10, respectively (p = 0.001). ELISA for human albumin in mice sera showed that levels were similar to those of control mice on day 2 (3.25 +/- 0.92 vs. 2.84 +/- 0.59 ng/ml, mean +/- SEM), peaked at 18.04 +/- 3.11 ng/ml on day 7, and decreased to 8.93 +/- 1.40 and 3.54 +/- 0.87 ng/ml on day 14 and 21, respectively (p = 0.02). Real-time quantitative RT-PCR showed gene expression levels of human albumin, alpha1-antitrypsin, and transferrin in mouse liver were 60.7 +/- 6.5%, 26.0 +/- 1.4%, and 156.8 +/- 62.4% of those of primary human adult hepatocytes, respectively, and immunohistochemistry revealed cells with human albumin and alpha1-antitrypsin expression in the mouse liver. In conclusion, our study demonstrated that bioluminescent imaging appears to be a sensitive, noninvasive modality for serial monitoring of transplanted hepatic stem cells.

  View details for Web of Science ID 000261489600003

  View details for PubMedID 19069633

• Molecular imaging of PET reporter gene expression. Handbook of experimental pharmacology Min, J., Gambhir, S. S. 2008: 277-303

  Abstract

  Multimodality molecular imaging continues to rapidly expand and is impacting many areas of biomedical research as well as patient management. Reporter-gene assays have emerged as a very general strategy for indirectly monitoring various intracellular events. Furthermore, reporter genes are being used to monitor gene/cell therapies, including the location(s), time variation, and magnitude of gene expression. This chapter reviews reporter gene technology and its major pre-clinical and clinical applications to date. The future appears quite promising for the continued expansion of the use of reporter genes in many evolving biomedically related arenas.

  View details for DOI 10.1007/978-3-540-77496-9_12

  View details for PubMedID 18626607

• Dual targeted contrast agent for US assessment of tumor angiogenesis in vivo Radiology Willmann J, Lutz A, Paulmurugan R, Patel M, Chu P, Rosenberg J, Gambhir SS 2008; 248 (3): 936-944
• Preclinical efficacy of the c-met inhibitor CE-355621 in a U87 MG mouse xenograft model evaluated by F-18-FDG small-animal PET JOURNAL OF NUCLEAR MEDICINE Tseng, J. R., Kang, K. W., Dandekar, M., Yaghoubi, S., Lee, J. H., Christensen, J. G., Muir, S., Vincent, P. W., Michaud, N. R., Gambhir, S. S. 2008; 49 (1): 129-134

  Abstract

  The purpose of this study was to evaluate the efficacy of CE-355621, a novel antibody against c-Met, in a subcutaneous U87 MG xenograft mouse model using (18)F-FDG small-animal PET.CE-355621 or control vehicle was administered intraperitoneally into nude mice (drug-treated group, n = 12; control group, n = 14) with U87 MG subcutaneous tumor xenografts. Drug efficacy was evaluated over 2 wk using (18)F-FDG small-animal PET and compared with tumor volume growth curves.The maximum %ID/g (percentage injected dose per gram of tissue) of (18)F-FDG accumulation in mice treated with CE-355621 remained essentially unchanged over 2 wk, whereas the %ID/g of the control tumors increased 66% compared with the baseline. Significant inhibition of (18)F-FDG accumulation was seen 3 d after drug treatment, which was earlier than the inhibition of tumor volume growth seen at 7 d after drug treatment.CE-355621 is an efficacious novel antineoplastic chemotherapeutic agent that inhibits (18)F-FDG accumulation earlier than tumor volume changes in a mouse xenograft model. These results support the use of (18)F-FDG PET to assess early tumor response for CE-355621.

  View details for DOI 10.2967/jnumed.106.038836

  View details for Web of Science ID 000252391700026

  View details for PubMedID 18077531

  View details for PubMedCentralID PMC4161137

• Molecular imaging of the efficacy of heat shock protein 90 inhibitors in living subjects CANCER RESEARCH Chan, C. T., Paulmurugan, R., Gheysens, O. S., Kim, J., Chiosis, G., Gambhir, S. S. 2008; 68 (1): 216-226

  Abstract

  Heat shock protein 90 alpha (Hsp90 alpha)/p23 and Hsp90 beta/p23 interactions are crucial for proper folding of proteins involved in cancer and neurodegenerative diseases. Small molecule Hsp90 inhibitors block Hsp90 alpha/p23 and Hsp90 beta/p23 interactions in part by preventing ATP binding to Hsp90. The importance of isoform-selective Hsp90 alpha/p23 and Hsp90 beta/p23 interactions in determining the sensitivity to Hsp90 was examined using 293T human kidney cancer cells stably expressing split Renilla luciferase (RL) reporters. Interactions between Hsp90 alpha/p23 and Hsp90 beta/p23 in the split RL reporters led to complementation of RL activity, which was determined by bioluminescence imaging of intact cells in cell culture and living mice using a cooled charge-coupled device camera. The three geldanamycin-based and seven purine-scaffold Hsp90 inhibitors led to different levels of inhibition of complemented RL activities (10-70%). However, there was no isoform selectivity to both classes of Hsp90 inhibitors in cell culture conditions. The most potent Hsp90 inhibitor, PU-H71, however, led to a 60% and 30% decrease in RL activity (14 hr) in 293T xenografts expressing Hsp90 alpha/p23 and Hsp90 beta/p23 split reporters respectively, relative to carrier control-treated mice. Molecular imaging of isoform-specific Hsp90 alpha/p23 and Hsp90 beta/p23 interactions and efficacy of different classes of Hsp90 inhibitors in living subjects have been achieved with a novel genetically encoded reporter gene strategy that should help in accelerating development of potent and isoform-selective Hsp90 inhibitors.

  View details for DOI 10.1158/0008-5472.CAN-07-2268

  View details for Web of Science ID 000252072100029

  View details for PubMedID 18172314

  View details for PubMedCentralID PMC4146344

• Molecular Imaging of Reporter Gene Expression in Prostate Cancer: An Overview. Seminars in Nuclear Medicine Singh A, Massoud T, Deroose C, Gambhir SS 2008; 39(1): 9-19
• I-123 MIBG mapping with intraoperative gamma probe for recurrent neuroblastoma MOLECULAR IMAGING AND BIOLOGY Iagaru, A., Peterson, D., Quon, A., Dutta, S., Twist, C., Daghighian, F., Gambhir, S. S., Albanese, C. 2008; 10 (1): 19-23

  Abstract

  Intraoperative gamma probe guidance has become widely utilized for sentinel lymph node dissection in patients with breast cancer and melanoma, using (99m)Tc sulfur colloid. However, new indications are possible and need to continue to be investigated. We report the use during a wedge liver biopsy of a new hand-held gamma probe designed for (123)I intraoperative guidance. The patient studied is a 5-year-old boy with history of stage 4 high-risk neuroblastoma. Anatomic imaging (CT, MRI), (99m)Tc bone scintigraphy and 2-deoxy-2-[F-18]fluoro-d-glucose-positron emission tomography/computed tomography (FDG-PET/CT) were negative, but the (123)I MIBG scintigraphy suggested recurrent liver disease. A decision was made to biopsy these lesions to obtain histopathological confirmation. Intraoperative gamma probe mapping of the liver identified areas with signal above the background, but these were prove to be hemosiderin deposits on histo-pathology examination.

  View details for DOI 10.1007/s11307-007-0116-1

  View details for Web of Science ID 000252107800002

  View details for PubMedID 17975716

• MOLECULAR IMAGING OF TUMOR VASCULATURE ANGIOGENESIS: IN VIVO SYSTEMS, PT B Cai, W., Gambhir, S. S., Chen, X. 2008; 445: 141-?

  Abstract

  Cancer, with more than 10 million new cases a year worldwide, is the third leading cause of death in developed countries. One critical requirement during cancer progression is angiogenesis, the formation of new blood vessels. Structural and functional imaging of tumor vasculature has been studied using various imaging modalities such as magnetic resonance imaging (MRI), computed tomography (CT), and ultrasound. Molecular imaging, a key component of the 21st-century cancer-patient management strategy, takes advantage of these traditional imaging techniques and introduces molecular probes to determine the expression of indicative molecular markers at different stages of cancer development. In this chapter, we will focus on two tumor vasculature-related targets: integrin alpha(v)beta(3) and vascular endothelial growth factor receptor (VEGFR). For imaging of integrin alpha(v)beta(3) on the tumor vasculature, only nanoparticle-based probes will be discussed. VEGFR imaging will be discussed in depth, and we will give a detailed example of positron emission tomography (PET) imaging of VEGFR expression using radio-labeled VEGF(121) protein. Future clinical translation will be critical for maximum patient benefit from these agents. To achieve this goal, multidisciplinary approaches and cooperative efforts from many individuals, institutions, industries, and organizations are needed to quickly translate multimodality tumor vasculature imaging into multiple facets of cancer patient management.

  View details for DOI 10.1016/S0076-6879(08)03007-3

  View details for Web of Science ID 000260774900007

  View details for PubMedID 19022059

• Applications of lentiviral vectors in non-invasive molecular imaging Methods in Molecular Biology De A, Yaghoubi S, Gambhir SS 2008; 433 (1): 177-202
• Ovarian cancer early detection claims are biased Clinical Center Researh McIntosh M, Anderson G, Drescher C, Hanash S, Urban N, Brown P, Gambhir SS, Coukos G, Laird P, Nelson B, Palmer C 2008; 14 (22): 7574
• Monitoring caspase-3 activation with a multimodality imaging sensor in living subjects Clinical Cancer Research Ray P, De A, Patel M, Gambhir SS 2008; 14 (18): 5801-5809
• Targeted microbubbles for imaging tumor angiogenesis: Assessment of whole body biodistribution with dynamic micro-PET in mice Radiology Willmann J, Cheng Z, Davis C, Lutz A, Schipper M, Nielsen C, Gambhir SS 2008; 249: 212-219
• Uptake Kinetics and Biodistribution of (14)C-D:-Luciferin-a Radiolabeled Substrate for the Firefly Luciferase Catalyzed Bioluminescence Reaction: Impact on Bioluminescence Based Reporter Gene Imaging European Journal of Nuclear Medicine and Molecular Imaging Berger, F., Paulmurugan, R, Bhaumik, S, Gambhir, SS 2008
• A comparison between a time domain and continuous wave small animal optical imaging system IEEE TRANSACTIONS ON MEDICAL IMAGING Keren, S., Gheysens, O., Levin, C. S., Gambhir, S. S. 2008; 27 (1): 58-63

  Abstract

  We present a phantom study to evaluate the performance of the eXplore Optix (Advanced Research Technologies-GE Healthcare), the first commercially available time-domain tomography system for small animal fluorescence imaging, and compare its capabilities with the widely used IVIS 200 (Xenogen Corporation-Caliper) continuous wave planar imaging system. The eXplore Optix, based on point-wise illumination and collection scheme, is found to be a log order more sensitive with significantly higher detection depth and spatial resolution as compared with the wide-area illumination IVIS 200 under the conditions tested. A time-resolved detection system allows the eXplore Optix to measure the arrival time distribution of fluorescence photons. This enables fluorescence lifetime measurement, absorption mapping, and estimation of fluorescent inclusion depth, which in turn is used by a reconstruction algorithm to calculate the volumetric distribution of the fluorophore concentration. An increased acquisition time and lack of ability to image multiple animals simultaneously are the main drawbacks of the eXplore Optix as compared with the IVIS 200.

  View details for DOI 10.1109/TMI.2007.902800

  View details for Web of Science ID 000252098300007

  View details for PubMedID 18270062

• BRET-Based method for detection of specific RNA species BIOCONJUGATE CHEMISTRY Walls, Z. F., Gambhir, S. S. 2008; 19 (1): 178-184

  Abstract

  RNA detection and quantitation is a common necessity in modern molecular biology research. Most methods, however, are complex and/or time-intensive. Presented here is a BRET (bioluminescene resonance energy transfer)-based method that can accomplish the task of RNA identification quickly and easily. By conjugating BRET enzymes to two different oligonucleotides that are complementary to the same target sequence, probes were developed that could detect RNA using a solution-based assay. This assay was optimized for spacer length between the binding sites (found to be 10 nucleotides), and sensitivity was determined to be 1 microg for a specific species of RNA within a mixed population. Specificity of the assay was assessed using in vitro transcribed cRNA and found to be statistically siginificant ( p = 3.11 x 10 (-6), ANOVA, multiple range test). This assay represents a possibility for a less technically demanding, streamlined alternative to canonical RNA assays.

  View details for DOI 10.1021/bc700278n

  View details for Web of Science ID 000252520300024

  View details for PubMedID 18072724

• Comparison between adenoviral and retroviral vectors for the transduction of the thymidine kinase PET reporter gene in rat mesenchymal stem cells Journal of Nuclear Medicine Roelants V, Labar D, DeMeester C, Havaux X, Tabilio A, Gambhir SS, Dilanni M, Bertrand L, Vanoverschelde J 2008; 49 (11): 1836-1844
• Crystal structures of the luciferase and green fluorescent protein from Renilla reniformis JOURNAL OF MOLECULAR BIOLOGY Loening, A. M., Fenn, T. D., Gambhir, S. S. 2007; 374 (4): 1017-1028

  Abstract

  Due to its ability to emit light, the luciferase from Renilla reniformis (RLuc) is widely employed in molecular biology as a reporter gene in cell culture experiments and small animal imaging. To accomplish this bioluminescence, the 37-kDa enzyme catalyzes the degradation of its substrate coelenterazine in the presence of molecular oxygen, resulting in the product coelenteramide, carbon dioxide, and the desired photon of light. We successfully crystallized a stabilized variant of this important protein (RLuc8) and herein present the first structures for any coelenterazine-using luciferase. These structures are based on high-resolution data measured to 1.4 A and demonstrate a classic alpha/beta-hydrolase fold. We also present data of a coelenteramide-bound luciferase and reason that this structure represents a secondary conformational form following shift of the product out of the primary active site. During the course of this work, the structure of the luciferase's accessory green fluorescent protein (RrGFP) was also determined and shown to be highly similar to that of Aequorea victoria GFP.

  View details for DOI 10.1016/j.jmb.2007.09.078

  View details for PubMedID 17980388

• Regulatory and reimbursement challenges for molecular imaging RADIOLOGY Hoffman, J. M., Gambhir, S. S., Kelloff, G. J. 2007; 245 (3): 645-660

  Abstract

  Molecular imaging is being hailed as the next great advance for imaging. Since molecular imaging typically involves the use of specific imaging probes that are treated like drugs, they will require regulatory approval. As with any drug, molecular imaging probes and techniques will also require thorough assessment in clinical trials to show safety and efficacy. The timeline for the regulatory approval will be long and potentially problematic because of the mounting costs of obtaining final regulatory approval. The current article is a detailed review of the regulatory and reimbursement process that will be required for molecular imaging probes and techniques to become a widespread clinical reality. The role of molecular imaging in the therapeutic drug discovery process will also be reviewed, as this is where these exciting new techniques have the potential to revolutionize the drug discovery and development process and, it is hoped, make it less costly. [(18)F]fluoro-2-deoxy-2-D-glucose positron emission tomography, one of the first molecular imaging techniques to be widely used, will be used as an example to illustrate the process of obtaining eventual reimbursement for widespread clinical use.

  View details for DOI 10.1148/radiol.2453060737

  View details for Web of Science ID 000251070700006

  View details for PubMedID 18024447

• Drug delivery - Keeping tabs on nanocarriers NATURE NANOTECHNOLOGY de la Zerda, A., Gambhir, S. S. 2007; 2 (12): 745-746

  View details for DOI 10.1038/nnano.2007.399

  View details for Web of Science ID 000251456500007

  View details for PubMedID 18654423

• Glia-dependent TGF-beta signaling, acting independently of the TH17 pathway, is critical for initiation of murine autoimmune encephalomyelitis JOURNAL OF CLINICAL INVESTIGATION Luo, J., Ho, P. P., Buckwalter, M. S., Hsu, T., Lee, L. Y., Zhang, H., Kim, D., Kim, S., Gambhir, S. S., Steinman, L., Wyss-Coray, T. 2007; 117 (11): 3306-3315

  Abstract

  Autoimmune encephalomyelitis, a mouse model for multiple sclerosis, is characterized by the activation of immune cells, demyelination of axons in the CNS, and paralysis. We found that TGF-beta1 synthesis in glial cells and TGF-beta-induced signaling in the CNS were activated several days before the onset of paralysis in mice with autoimmune encephalomyelitis. While early production of TGF-beta1 was observed in glial cells TGF-beta signaling was activated in neurons and later in infiltrating T cells in inflammatory lesions. Systemic treatment with a pharmacological inhibitor of TGF-beta signaling ameliorated the paralytic disease and reduced the accumulation of pathogenic T cells and expression of IL-6 in the CNS. Priming of peripheral T cells was not altered, nor was the generation of TH17 cells, indicating that this effect was directed within the brain, yet affected the immune system. These results suggest that early production of TGF-beta1 in the CNS creates a permissive and dangerous environment for the initiation of autoimmune inflammation, providing a rare example of the brain modulating the immune system. Importantly, inhibition of TGF-beta signaling may have benefits in the treatment of the acute phase of autoimmune CNS inflammation.

  View details for DOI 10.1172/JCI31763

  View details for PubMedID 17965773

• Dual-function probe for PET and near-infrared fluorescence imaging of tumor vasculature JOURNAL OF NUCLEAR MEDICINE Cai, W., Chen, K., Li, Z., Gambhir, S. S., Chen, X. 2007; 48 (11): 1862-1870

  Abstract

  To date, the in vivo imaging of quantum dots (QDs) has been mostly qualitative or semiquantitative. The development of a dual-function PET/near-infrared fluorescence (NIRF) probe can allow for accurate assessment of the pharmacokinetics and tumor-targeting efficacy of QDs.A QD with an amine-functionalized surface was modified with RGD peptides and 1,4,7,10-tetraazacyclodocecane-N,N',N'',N'''-tetraacetic acid (DOTA) chelators for integrin alpha(v)beta(3)-targeted PET/NIRF imaging. A cell-binding assay and fluorescence cell staining were performed with U87MG human glioblastoma cells (integrin alpha(v)beta(3)-positive). PET/NIRF imaging, tissue homogenate fluorescence measurement, and immunofluorescence staining were performed with U87MG tumor-bearing mice to quantify the probe uptake in the tumor and major organs.There are about 90 RGD peptides per QD particle, and DOTA-QD-RGD exhibited integrin alpha(v)beta(3)-specific binding in cell cultures. The U87MG tumor uptake of (64)Cu-labeled DOTA-QD was less than 1 percentage injected dose per gram (%ID/g), significantly lower than that of (64)Cu-labeled DOTA-QD-RGD (2.2 +/- 0.3 [mean +/- SD] and 4.0 +/- 1.0 %ID/g at 5 and 18 h after injection, respectively; n = 3). Taking into account all measurements, the liver-, spleen-, and kidney-to-muscle ratios for (64)Cu-labeled DOTA-QD-RGD were about 100:1, 40:1, and 1:1, respectively. On the basis of the PET results, the U87MG tumor-to-muscle ratios for DOTA-QD-RGD and DOTA-QD were about 4:1 and 1:1, respectively. Excellent linear correlation was obtained between the results measured by in vivo PET imaging and those measured by ex vivo NIRF imaging and tissue homogenate fluorescence (r(2) = 0.93). Histologic examination revealed that DOTA-QD-RGD targets primarily the tumor vasculature through an RGD-integrin alpha(v)beta(3) interaction, with little extravasation.We quantitatively evaluated the tumor-targeting efficacy of a dual-function QD-based probe with PET and NIRF imaging. This dual-function probe has significantly reduced potential toxicity and overcomes the tissue penetration limitation of optical imaging, allowing for quantitative targeted imaging in deep tissue.

  View details for DOI 10.2967/jnumed.107.043216

  View details for Web of Science ID 000252894900024

  View details for PubMedID 17942800

• Molecular imaging techniques in body imaging RADIOLOGY Margolis, D. J., Hoffman, J. M., Herfkens, R. J., Jeffrey, R. B., Quon, A., Gambhir, S. S. 2007; 245 (2): 333-356

  Abstract

  Molecular imaging of the body involves new techniques to image cellular biochemical processes, which results in studies with high sensitivity, specificity, and signal-to-background. The most prevalently used molecular imaging technique in body imaging is currently fluorine 18 fluorodeoxyglucose (FDG) positron emission tomography (PET). FDG PET has become the method of choice for the staging and restaging of many of the most common cancers, including lymphoma, lung cancer, breast cancer, and colorectal cancer. FDG PET has also become extremely valuable in monitoring the response to therapeutic drugs in many cancers. New PET agents, such as fluorothymidine and acetate, have also shown promise in the evaluation of response to therapy and in the staging of prostate cancer. Magnetic resonance (MR) spectroscopy has shown promise in the evaluation of prostate cancer. Breast cancer evaluation benefits from advances in spectroscopic imaging and contrast-enhanced kinetic evaluation of vascular permeability, which is altered in neoplastic processes because of release of angiogenic factors. Superparamagnetic iron oxide (SPIO) particles represent the first of an expanding line of MR contrast agents that target specific cellular processes. SPIO particles have also been used in the evaluation of the cirrhotic liver and at MR lymphangiography.

  View details for DOI 10.1148/radiol.2452061117

  View details for Web of Science ID 000250343800007

  View details for PubMedID 17940297

• Multimodality imaging of T-cell hybridoma trafficking in collagen-induced arthritic mice: image-based estimation of the number of cells accumulating in mouse paws JOURNAL OF BIOMEDICAL OPTICS Yaghoubi, S. S., Creusot, R. J., Ray, P., Fathman, C. G., Gambhir, S. S. 2007; 12 (6)

  Abstract

  Appropriate targeting of therapeutic cells is essential in adoptive cellular gene therapy (ACGT). Imaging cell trafficking in animal models and patients will guide development of ACGT protocols. Collagen type II (C-II)-specific T cell hybridomas are transduced with a lentivirus carrying a triple fusion reporter gene (TFR) construct consisting of a fluorescent reporter gene (RG), a bioluminescent RG (hRluc), and a positron emission tomography (PET) RG. Collagen-induced arthritic (CIA) mice are scanned with a bioluminescence imaging camera before and after implantation of various known cell quantities in their paws. Linear regression analysis yields equations relating two parameters of image signal intensity in mice paws to the quantity of hRluc expressing cells in the paws. Afterward, trafficking of intravenously injected cells is studied by quantitative analysis of bioluminescence images. Comparison of the average cell numbers does not demonstrate consistently higher accumulation of T-cell hybridomas in the paws with higher inflammation scores, and injecting more cells does not cause increased accumulation. MicroPET images illustrate above background signal in the inflamed paws and chest areas of CIA mice. The procedures described in this study can be used to derive equations for cells expressing other bioluminescent RGs and in other animal models.

  View details for DOI 10.1117/1.2821415

  View details for Web of Science ID 000252851100046

  View details for PubMedID 18163841

• Design and evaluation of a variable aperture collimator for conformal radiotherapy of small animals using a microCT scanner MEDICAL PHYSICS Graves, E. E., Zhou, H., Chatterjee, R., Keall, P. J., Gambhir, S. S., Contag, C. H., Boyer, A. L. 2007; 34 (11): 4359-4367

  Abstract

  Treatment of small animals with radiation has in general been limited to planar fields shaped with lead blocks, complicating spatial localization of dose and treatment of deep-seated targets. In order to advance laboratory radiotherapy toward what is accomplished in the clinic, we have constructed a variable aperture collimator for use in shaping the beam of microCT scanner. This unit can image small animal subjects at high resolution, and is capable of delivering therapeutic doses in reasonable exposure times. The proposed collimator consists of two stages, each containing six trapezoidal brass blocks that move along a frame in a manner similar to a camera iris producing a hexagonal aperture of variable size. The two stages are offset by 30 degrees and adjusted for the divergence of the x-ray beam so as to produce a dodecagonal profile at isocenter. Slotted rotating driving plates are used to apply force to pins in the collimator blocks and effect collimator motion. This device has been investigated through both simulation and measurement. The collimator aperture size varied from 0 to 8.5 cm as the driving plate angle increased from 0 to 41 degrees. The torque required to adjust the collimator varied from 0.5 to 5 N x m, increasing with increasing driving plate angle. The transmission profiles produced by the scanner at isocenter exhibited a penumbra of approximately 10% of the collimator aperture width. Misalignment between the collimator assembly and the x-ray source could be identified on the transmission images and corrected by adjustment of the collimator location. This variable aperture collimator technology is therefore a feasible and flexible solution for adjustable shaping of radiation beams for use in small animal radiotherapy as well as other applications in which beam shaping is desired.

  View details for DOI 10.1118/1.2789498

  View details for Web of Science ID 000251145900029

  View details for PubMedID 18072501

• Invasive assessment of coronary physiology using a pressure wire correlates with cardiac positron emission tomography in patients with coronary disease. Brinton, T. J., Quon, A., Schindler, T. H., Luero, J. L., Hansen, H., Segall, G., Giacormni, J. C., Schellbert, H. R., Gambhir, S. S., Fearon, W. F. LIPPINCOTT WILLIAMS & WILKINS. 2007: 751

  View details for Web of Science ID 000250394303430

• Quantum dot imaging for embryonic stem cells BMC BIOTECHNOLOGY Lin, S., Xie, X., Patel, M. R., Yang, Y., Li, Z., Cao, F., Gheysens, O., Zhang, Y., Gambhir, S. S., Rao, J. H., Wu, J. C. 2007; 7

  Abstract

  Semiconductor quantum dots (QDs) hold increasing potential for cellular imaging both in vitro and in vivo. In this report, we aimed to evaluate in vivo multiplex imaging of mouse embryonic stem (ES) cells labeled with Qtracker delivered quantum dots (QDs).Murine embryonic stem (ES) cells were labeled with six different QDs using Qtracker. ES cell viability, proliferation, and differentiation were not adversely affected by QDs compared with non-labeled control cells (P = NS). Afterward, labeled ES cells were injected subcutaneously onto the backs of athymic nude mice. These labeled ES cells could be imaged with good contrast with one single excitation wavelength. With the same excitation wavelength, the signal intensity, defined as (total signal-background)/exposure time in millisecond was 11 +/- 2 for cells labeled with QD 525, 12 +/- 9 for QD 565, 176 +/- 81 for QD 605, 176 +/- 136 for QD 655, 167 +/- 104 for QD 705, and 1,713 +/- 482 for QD 800. Finally, we have shown that QD 800 offers greater fluorescent intensity than the other QDs tested.In summary, this is the first demonstration of in vivo multiplex imaging of mouse ES cells labeled QDs. Upon further improvements, QDs will have a greater potential for tracking stem cells within deep tissues. These results provide a promising tool for imaging stem cell therapy non-invasively in vivo.

  View details for DOI 10.1186/1472-6750-7-67

  View details for Web of Science ID 000252448600001

  View details for PubMedID 17925032

  View details for PubMedCentralID PMC2174930

• Bisdeoxycoelenterazine derivatives for improvement of bioluminescence resonance energy transfer assays JOURNAL OF THE AMERICAN CHEMICAL SOCIETY Levi, J., De, A., Cheng, Z., Gambhir, S. S. 2007; 129 (39): 11900-?

  View details for DOI 10.1021/ja073936h

  View details for Web of Science ID 000249887700013

  View details for PubMedID 17850082

• Molecular imaging can accelerate anti-angiogenic drug development and testing NATURE CLINICAL PRACTICE ONCOLOGY Lagaru, A., Chen, X., Gambhir, S. S. 2007; 4 (10): 556-557

  View details for DOI 10.1038/ncponc0929

  View details for Web of Science ID 000249708800002

  View details for PubMedID 17726490

• Osseous and soft tissue sarcomas: When can F-18 FDG PET/CT evaluation provide useful information? 20th Annual Congress of the European-Association-of-Nuclear-Medicine Iagaru, A., Quon, A., Jacobs, C., Marina, N., McDougall, I., Gambhir, S. S. SPRINGER. 2007: S152–S152

  View details for Web of Science ID 000253283900155

• Differentiation, survival, and function of embryonic stem cell-derived endothelial cells for ischemic heart disease 79th Annual Scientific Session of the American-Heart-Association Li, Z., Wu, J. C., Sheikh, A. Y., Kraft, D., Cao, F., Xie, X., Patel, M., Gambhir, S. S., Robbins, R. C., Cooke, J. P., Wu, J. C. LIPPINCOTT WILLIAMS & WILKINS. 2007: I46–I54

  Abstract

  Embryonic stem (ES) cells are distinguished by their capacity for self-renewal and pluripotency. Here we characterize the differentiation of ES cell-derived endothelial cells (ESC-ECs), use molecular imaging techniques to examine their survival in vivo, and determine the therapeutic efficacy of ESC-ECs for restoration of cardiac function after ischemic injury.Murine ES cells were transfected with a construct composed of a vascular endothelial cadherin promoter driving enhanced green fluorescence protein (pVE-cadherin-eGFP). Differentiation of ES cells to ECs was detected by FACS analysis using Flk-1 (early EC marker at day 4) and VE-cadherin (late EC marker at day 8). After isolation, these ESC-ECs express endothelial cell markers similar to adult mouse lung endothelial cells, form vascular-like channels, and incorporate DiI-labeled acetylated low-density lipoprotein (DiI-Ac-LDL). For in vivo imaging, ES cells were transduced with an ubiquitin promoter driving firefly luciferase and monomeric red fluorescence protein (pUb-Fluc-mRFP). A robust correlation exists between Fluc signals and cell numbers by ex vivo imaging analysis (R2=0.98) and by in vitro enzyme assay (R2=0.94). Afterward, 5x10(5) ESC-ECs or PBS (as control) was injected into the hearts of mice undergoing LAD ligation (n=15 per group). Bioluminescence imaging showed longitudinal survival of transplanted ESC-ECs for approximately 8 weeks. Echocardiogram demonstrated significant functional improvement in the ESC-EC group compared with control (P=0.04). Finally, postmortem analysis confirmed increased presence of small capillaries and venules in the infarcted zones by CD31 staining.This is the first study to track the fate and function of transplanted ESC-ECs in the heart. With further validation, these ESC-ECs could become a valuable source of cell therapy for induction of angiogenesis in the treatment of myocardial ischemia.

  View details for DOI 10.1161/CIRCULATIONAHA.106.680561

  View details for PubMedID 17846325

• Molecular imaging: Integration of molecular imaging into the musculoskeletal imaging practice RADIOLOGY Biswal, S., Resnick, D. L., Hoffman, J. M., Gambhir, S. S. 2007; 244 (3): 651-671

  Abstract

  Chronic musculoskeletal diseases such as arthritis, malignancy, and chronic injury and/or inflammation, all of which may produce chronic musculoskeletal pain, often pose challenges for current clinical imaging methods. The ability to distinguish an acute flare from chronic changes in rheumatoid arthritis, to survey early articular cartilage breakdown, to distinguish sarcomatous recurrence from posttherapeutic inflammation, and to directly identify generators of chronic pain are a few examples of current diagnostic limitations. There is hope that a growing field known as molecular imaging will provide solutions to these diagnostic puzzles. These techniques aim to depict, noninvasively, specific abnormal cellular, molecular, and physiologic events associated with these and other diseases. For example, the presence and mobilization of specific cell populations can be monitored with molecular imaging. Cellular metabolism, stress, and apoptosis can also be followed. Furthermore, disease-specific molecules can be targeted, and particular gene-related events can be assayed in living subjects. Relatively recent molecular and cellular imaging protocols confirm important advances in imaging technology, engineering, chemistry, molecular biology, and genetics that have coalesced into a multidisciplinary and multimodality effort. Molecular probes are currently being developed not only for radionuclide-based techniques but also for magnetic resonance (MR) imaging, MR spectroscopy, ultrasonography, and the emerging field of optical imaging. Furthermore, molecular imaging is facilitating the development of molecular therapies and gene therapy, because molecular imaging makes it possible to noninvasively track and monitor targeted molecular therapies. Implementation of molecular imaging procedures will be essential to a clinical imaging practice. With this in mind, the goal of the following discussion is to promote a better understanding of how such procedures may help address specific musculoskeletal issues, both now and in the years ahead.

  View details for DOI 10.1148/radiol.2443060295

  View details for Web of Science ID 000248993500006

  View details for PubMedID 17709823

• Fusion of Gaussia luciferase to an engineered anti-carcinoembryonic antigen (CEA) antibody for in vivo optical imaging MOLECULAR IMAGING AND BIOLOGY Venisnik, K. M., Olafsen, T., Gambhir, S. S., Wu, A. M. 2007; 9 (5): 267-277

  Abstract

  The bioluminescent protein Gaussia luciferase (GLuc) was fused to an anti-carcinoembryonic antigen (CEA) antibody fragment, the diabody, for in vivo optical tumor imaging. A 15-amino acid N-terminal truncation (GLDelta15) resulted in a brighter protein. Fusions of the anti-CEA diabody to full-length GLuc and GLDelta15 retained high affinity for the antigen, emitted light, and exhibited excellent enzymatic stability. In vivo optical imaging of tumor-bearing mice demonstrated specific targeting of diabody-GLDelta15 to CEA-positive xenografts, with a tumor/background ratio of 3.8 +/- 0.4 at four hours after tail-vein injection, compared to antigen-negative tumors at 1.3 +/- 0.1 (p = 0.001). MicroPET imaging using (124)I-diabody-GLDelta15 demonstrated specific uptake in the CEA-positive tumor (2.6% ID [injected dose]/g) compared to the CEA-negative tumor (0.4% ID/g) at 21 hours. Although further optimization of this fusion protein may be needed to improve in vivo performance, the diabody-GLDelta15 is a promising optical imaging probe for tumor detection in vivo.

  View details for DOI 10.1007/s11307-007-0101-8

  View details for Web of Science ID 000248865200003

  View details for PubMedID 17577599

• MicroPET-based biodistribution of quantum dots in living mice JOURNAL OF NUCLEAR MEDICINE Schipper, M. L., Cheng, Z., Lee, S., Bentolila, L. A., Iyer, G., Rao, J., Chen, X., Wu, A. M., Weiss, S., Gambhir, S. S. 2007; 48 (9): 1511-1518

  Abstract

  This study evaluates the quantitative biodistribution of commercially available CdSe quantum dots (QD) in mice.(64)Cu-Labeled 800- or 525-nm emission wavelength QD (21- or 12-nm diameter), with or without 2,000 MW (molecular weight) polyethylene glycol (PEG), were injected intravenously into mice (5.55 MBq/25 pmol QD) and studied using well counting or by serial microPET and region-of-interest analysis.Both methods show rapid uptake by the liver (27.4-38.9 %ID/g) (%ID/g is percentage injected dose per gram tissue) and spleen (8.0-12.4 %ID/g). Size has no influence on biodistribution within the range tested here. Pegylated QD have slightly slower uptake into liver and spleen (6 vs. 2 min) and show additional low-level bone uptake (6.5-6.9 %ID/g). No evidence of clearance from these organs was observed.Rapid reticuloendothelial system clearance of QD will require modification of QD for optimal utility in imaging living subjects. Formal quantitative biodistribution/imaging studies will be helpful in studying many types of nanoparticles, including quantum dots.

  View details for DOI 10.2967/jnumed.107.040071

  View details for PubMedID 17704240

• 2-Deoxy-2-[F-18]fluoro-D-glucose accumulation in ovarian carcinoma cell lines MOLECULAR IMAGING AND BIOLOGY Lutz, A. M., Ray, P., Willmann, J. K., Drescher, C., Gambhir, S. S. 2007; 9 (5): 260-266

  Abstract

  To evaluate 2-deoxy-2-[F-18]fluoro-D-glucose (FDG) accumulation in human ovarian carcinoma cell lines compared with control tumor cell lines known to accumulate FDG.FDG accumulation assays were performed in 15 different ovarian carcinoma cell lines at 1, 2, and 3 hours after incubation with 1 microCi of FDG. Results were compared with FDG accumulation in six different control tumor cell lines. 2-deoxy-2-[F-18]fluoro-D-glucose accumulation was expressed as counts per minute (cpm) in cells and normalized to initial cpm in medium and total protein content of cell lysates.FDG accumulation in all 15 ovarian carcinoma cell lines was equal to or higher than 0.0005 +/- 8.6 10(-5) cpm in cells/cpm in medium/mug protein at all three different time points. In two ovarian carcinoma cell lines (ES-2, poorly differentiated clear cell carcinoma, and OVCAR-3, poorly differentiated papillary adenocarcinoma), FDG accumulation was not statistically, significantly different compared to the control cell line with the highest FDG accumulation (LS 174T human colorectal adenocarcinoma) at two or more time points (P > or = 0.07). In 2 of 15 (13%) ovarian carcinoma cell lines (OVCAR5 epithelial carcinoma and SKOV3 clear cell carcinoma), FDG accumulation was lower than that in the control cell line with the lowest FDG accumulation (HT-29 human colorectal adenocarcinoma) at one or more time points (P < 0.05).Most human ovarian carcinoma cell lines showed comparable FDG accumulations with control cell lines known to accumulate FDG. This study lays the foundations for further comparisons with other ovarian cancer cell lines and for other positron emission tomography tracers.

  View details for DOI 10.1007/s11307-007-0105-4

  View details for Web of Science ID 000248865200002

  View details for PubMedID 17610017

• An improved bioluminescence resonance energy transfer strategy for imaging intracellular events in single cells and living subjects CANCER RESEARCH De, A., Loening, A. M., Gambhir, S. S. 2007; 67 (15): 7175-7183

  Abstract

  Bioluminescence resonance energy transfer (BRET) is currently used for monitoring various intracellular events, including protein-protein interactions, in normal and aberrant signal transduction pathways. However, the BRET vectors currently used lack adequate sensitivity for imaging events of interest from both single living cells and small living subjects. Taking advantage of the critical relationship of BRET efficiency and donor quantum efficiency, we report generation of a novel BRET vector by fusing a GFP(2) acceptor protein with a novel mutant Renilla luciferase donor selected for higher quantum yield. This new BRET vector shows an overall 5.5-fold improvement in the BRET ratio, thereby greatly enhancing the dynamic range of the BRET signal. This new BRET strategy provides a unique platform to assay protein functions from both single live cells and cells located deep within small living subjects. The imaging utility of the new BRET vector is shown by constructing a sensor using two mammalian target of rapamycin pathway proteins (FKBP12 and FRB) that dimerize only in the presence of rapamycin. This new BRET vector should facilitate high-throughput sensitive BRET assays, including studies in single live cells and small living subjects. Applications will include anticancer therapy screening in cell culture and in small living animals.

  View details for DOI 10.1158/0008-5472.CAN-06-4623

  View details for PubMedID 17671185

• Red-shifted Renilla reniformis luciferase variants for imaging in living subjects NATURE METHODS Loening, A. M., Wu, A. M., Gambhir, S. S. 2007; 4 (8): 641-643

  Abstract

  The use of R. reniformis luciferase (RLuc) as a reporter gene in small-animal imaging has been hampered by its 481 nm peaked emission spectrum, as blue wavelengths are strongly attenuated in biological tissues. To overcome this, we generated variants of RLuc with bathochromic (red) shifts of up to 66 nm (547 nm peak) that also had greater stability and higher light emission than native RLuc.

  View details for DOI 10.1038/NMETH1070

  View details for PubMedID 17618292

• Cardiovascular molecular imaging RADIOLOGY Wu, J. C., Bengel, F. M., Gambhir, S. S. 2007; 244 (2): 337-355

  Abstract

  The goal of this review is to highlight how molecular imaging will impact the management and improved understanding of the major cardiovascular diseases that have substantial clinical impact and research interest. These topics include atherosclerosis, myocardial ischemia, myocardial viability, heart failure, gene therapy, and stem cell transplantation. Traditional methods of evaluation for these diseases will be presented first, followed by methods that incorporate conventional and molecular imaging approaches.

  View details for Web of Science ID 000248821400005

  View details for PubMedID 17592037

• Oxygen sensitivity of reporter genes: Implications for preclinical imaging of tumor hypoxia MOLECULAR IMAGING Cecic, I., Chan, D. A., Sutphin, P. D., Ray, P., Gambhir, S. S., Giaccia, A. J., Gravcs, E. E. 2007; 6 (4): 219-228

  Abstract

  Reporter gene techniques have been applied toward studying the physiologic phenomena associated with tumor hypoxia, a negative prognostic indicator. The purpose of this study was to assess the potential adverse effects of hypoxic conditions on the effectiveness of four commonly used reporter genes: Renilla luciferase, monomeric red fluorescent protein, thymidine kinase, and lacZ. Tumor-forming A375 cells expressing a trifusion reporter consisting of Renilla luciferase, monomeric red fluorescent protein, and thymidine kinase were subjected to decreasing oxygen tensions and assayed for reporter expression and activity. A375 cells expressing beta-galactosidase were similarly exposed to hypoxia, with activity of the reporter monitored by cleavage of the fluorescent substrate 7-hydroxy-9H-(1,3-dichloro-9,9-dimethylacridin-2-one)-beta-galactoside (DDAOG). Generation of signal in in vivo tumor models expressing bioluminescent or beta-galactosidase reporters were also examined over the course of hypoxic stresses, either by tumor clamping or the antivascular agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA). Our findings indicate that bioluminescent and fluorescent reporter activity are decreased under hypoxia despite minimal variations in protein production, whereas beta-galactosidase reporter activity per unit protein was unchanged. These results demonstrate that combining beta-galactosidase with the DDAOG optical probe may be a robust reporter system for the in vivo study of tumor hypoxia.

  View details for DOI 10.2310/7290.2007.00017

  View details for Web of Science ID 000249349100001

  View details for PubMedID 17711777

• Molecular imaging: The vision and opportunity for radiology in the future RADIOLOGY Hoffman, J. M., Gambhir, S. S. 2007; 244 (1): 39-47

  Abstract

  Molecular imaging is being hailed as the next great advance for imaging. This introductory article in the molecular imaging series to be published over the next several months in Radiology sets the stage for the subsequent set of articles by providing relevant definitions and background information and traces the evolution of molecular imaging to its current state of research and clinical practice. It discusses in detail the evolution of molecular imaging and the role that the National Cancer Institute and the National Institutes of Health have had in the funding and development of many of the important molecular imaging research programs that are in existence today. The article also provides basic information about the complex biology of the cell and details of the pathogenesis of cancer and how molecular imaging will be critical for earlier detection and management of cancer in the future. The article lays the foundation for the subsequent articles in the series and describes how and why molecular imaging will be critical and integral for clinical care of patients in the future. The introductory article also discusses the relevance of molecular imaging to clinical radiology practice and why it is critical for the practicing radiologist to understand these evolving techniques, as they will be the future of imaging.

  View details for DOI 10.1148/radiol.2441060773

  View details for Web of Science ID 000247436500005

  View details for PubMedID 17507723

• Small-animal PET of melanocortin 1 receptor expression using a F-18-labeled alpha-melanocyte-stimulating hormone analog JOURNAL OF NUCLEAR MEDICINE Cheng, Z., Zhang, L., Graves, E., Xiong, Z., Dandekar, M., Chen, X., Gambhir, S. S. 2007; 48 (6): 987-994

  Abstract

  (18)F-Labeled small synthetic peptides have emerged as attractive probes for imaging various molecular targets with PET. The alpha-melanocyte-stimulating hormone (alpha-MSH) receptor (melanocortin type 1 receptor [MC1R]) is overexpressed in most murine and human melanomas. It is a promising molecular target for diagnosis and therapy of melanomas. However, (18)F compounds have not been successfully developed for imaging the MC1R.In this study, an alpha-MSH analog, Ac-Nle-Asp-His-D-Phe-Arg-Trp-Gly-Lys-NH(2) (NAPamide), was radiolabeled with N-succinimidyl-4-(18)F-fluorobenzoate ((18)F-SFB). The resulting radiopeptide was evaluated as a potential molecular probe for small-animal PET of melanoma and MC1R expression in melanoma xenografted mouse models.The binding affinity of (19)F-SFB-conjugated NAPamide, (19)F-FB-NAPamide, was determined to be 7.2 +/- 1.2 nM (mean +/- SD) using B16/F10 cells and (125)I-(Tyr(2))-[Nle(4),D-Phe(7)]-alpha-MSH [(125)I-(Tyr(2))-NDP] as a radioligand. The biodistribution of (18)F-FB-NAPamide was then investigated in C57BL/6 mice bearing subcutaneous murine B16/F10 melanoma tumors with high expression of MC1Rs and Fox Chase Scid mice bearing human A375M melanoma with a relatively low number of MC1R receptors. Biodistribution experiments showed that tumor uptake values (percentage injected dose per gram of tumor [%ID/g]) of (18)F-FB-NAPamide were 1.19 +/- 0.11 %ID/g and 0.46 +/- 0.11 %ID/g, in B16/F10 and A375M xenografted melanoma at 1 h after injection, respectively. Furthermore, the B16/F10 tumor uptake was significantly inhibited by coinjection with excess alpha-MSH peptide (P < 0.05), indicating that (18)F-FB-NAPamide specifically recognizes the MC1R in living mice. Small-animal PET of (18)F-FB-NAPamide in mice bearing B16/F10 and A375M tumors at 1 h after tail vein injection revealed good B16/F10 tumor-to-background contrast and low A375M tumor-to-background ratios.(18)F-FB-NAPamide is a promising molecular probe for alpha-MSH receptor-positive melanoma PET and warrants further study.

  View details for DOI 10.2967/jnumed.107.039602

  View details for Web of Science ID 000247054800024

  View details for PubMedID 17504880

• Development of a variable-aperture collimator for small animal radiation Zhou, H., Chatterjee, R., Contag, C., Gambhir, S., Boyer, A., Keall, P., Graves, E. AMER ASSOC PHYSICISTS MEDICINE AMER INST PHYSICS. 2007: 2436

  View details for DOI 10.1118/1.2760811

  View details for Web of Science ID 000247479600511

• Evaluation of herpes simplex virus 1 thymidine kinase-mediated trapping of I-131 FIAU and prodrug activation of ganciclovir as a synergistic cancer radio/chemotherapy MOLECULAR IMAGING AND BIOLOGY Schipper, M. L., Goris, M. L., Gambhir, S. S. 2007; 9 (3): 110-116

  Abstract

  Evaluation of selective killing of Herpes Simplex Virus 1 thymidine kinase (HSV1-tk) expressing tumors by radiolabeled (131)I-fialuridine (FIAU), and of synergy between (131)I-FIAU and Ganciclovir (GCV).HSV1-tk-expressing cell lines and parental cell lines were exposed to (131)I-FIAU alone, GCV alone, or combinations. Activity and concentration were varied widely, concurrent and sequential administrations tested, and dose rate effects were studied.HSV1-tk-expressing cells accumulated up to 15.7-fold more (131)I-FIAU, were growth inhibited by 2 muCi/ml, or 5 muCi/ml (131)I-FIAU, and were inhibited by two log orders lower concentrations of GCV than parental cells. However, no synergy or additive effect was observed. Dose rate variations, or sequential treatment, did not alter outcome.Radioisotope therapy of HSV1-tk-expressing tumor cells with (131)I-FIAU is reported for the first time. Lack of synergy between (131)I-FIAU and GCV does not warrant further investigation of combination treatment with the two agents.

  View details for DOI 10.1007/s11307-007-0078-3

  View details for Web of Science ID 000246175500003

  View details for PubMedID 17294333

• Integrating noninvasive molecular imaging into molecular medicine: an evolving paradigm TRENDS IN MOLECULAR MEDICINE Massoud, T. F., Gambhir, S. S. 2007; 13 (5): 183-191

  Abstract

  Molecular imaging is a rapidly emerging field, providing noninvasive visual quantitative representations of fundamental biological processes in intact living subjects. Fundamental biomedical research stands to benefit considerably from advances in molecular imaging, with improved molecular target selection, probe development and imaging instrumentation. The noninvasiveness of molecular imaging technologies will also provide benefit through improved patient care. Molecular imaging endpoints can be quantified, and therefore are particularly useful for translational research. Integration of the two disciplines of molecular imaging and molecular medicine, combined with systems-biology approaches to understanding disease complexity, promises to provide predictive, preventative and personalized medicine that will transform healthcare.

  View details for DOI 10.1016/j.molmed.2007.03.003

  View details for Web of Science ID 000247166100002

  View details for PubMedID 17403616

• Fluorescent fructose derivatives for imaging breast cancer cells BIOCONJUGATE CHEMISTRY Levi, J., Cheng, Z., Gheysens, O., Patel, M., Chan, C. T., Wang, Y., Namavari, M., Gambhir, S. S. 2007; 18 (3): 628-634

  Abstract

  Breast cancer cells are known to overexpress Glut5, a sugar transporter responsible for the transfer of fructose across the cell membrane. Since Glut5 transporter is not significantly expressed in normal breast cells, fructose uptake can potentially be used to differentiate between normal and cancerous cells. Fructose was labeled with two fluorophores at the C-1 position: 7-nitro-1,2,3-benzadiazole (NBD) and Cy5.5. The labeling site was chosen on the basis of the presence and substrate specificity of the key proteins involved in the first steps of fructose metabolism. Using fluorescence microscopy, the uptake of the probes was studied in three breast cancer cell lines: MCF 7, MDA-MB-435, and MDA-MB-231. Both fluorescent fructose derivatives showed a very good uptake in all tested cell lines. The level of uptake was comparable to that of the corresponding glucose analogs, 2-NBDG and Cy5.5-DG. Significant uptake of 1-NBDF derivative was not observed in cells lacking Glut5 transporter, while the uptake of the 1-Cy5.5-DF derivative was independent of the presence of a fructose-specific transporter. While 1-NBDF showed Glut5-specific accumulation, the coupling of a large fluorophore such as Cy5.5 likely introduces big structural and electronic changes, leading to a fructose derivative that does not accurately describe the uptake of fructose in cells.

  View details for DOI 10.1021/bc060184s

  View details for Web of Science ID 000246485500005

  View details for PubMedID 17444608

  View details for PubMedCentralID PMC4145876

• Cu-64-Labeled alpha-melanocyte-stimulating hormone analog for MicroPET imaging of melanocortin 1 receptor expression BIOCONJUGATE CHEMISTRY Cheng, Z., Xiong, Z., Subbarayan, M., Chen, X., Gambhir, S. S. 2007; 18 (3): 765-772

  Abstract

  The alpha-melanocyte-stimulating hormone (alpha-MSH) receptor (melanocortin type 1 receptor, or MC1R) plays an important role in the development and growth of melanoma cells. It was found that MC1R was overexpressed on most murine and human melanoma, making it a promising molecular target for melanoma imaging and therapy. Radiolabeled alpha-MSH peptide and its analogs that can specifically bind with MC1R have been extensively explored for developing novel agents for melanoma detection and radionuclide therapy. The goal of this study was to evaluate a 64Cu-labeled alpha-MSH analog, Ac-Nle-Asp-His-D-Phe-Arg-Trp-Gly-Lys(DOTA)-NH2 (DOTA-NAPamide), as a potential molecular probe for microPET imaging of melanoma and MC1R expression in melanoma xenografted mouse models. 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) conjugated NAPamide was synthesized and radiolabeled with 64Cu (t1/2=12 h) in NH4OAc (0.1 M; pH 5.5) buffered solution for 60 min at 50 degrees C. Cell culture studies reveal rapid and high uptake and internalization of 64Cu-DOTA-NAPamide in B16F10 cells. Over 90% of receptor-bound tracer is internalized at 3 h incubation. A cellular retention study demonstrates that the receptor-bound 64Cu-DOTA-NAPamide is slowly released from the B16F10 cells into the medium; 66% of the radioactivity is still associated with the cells even after 3 h incubation. The biodistribution of 64Cu-DOTA-NAPamide was then investigated in C57BL/6 mice bearing subcutaneous murine B16F10 melanoma tumors with high capacity of MC1R and Fox Chase Scid mice bearing human A375M melanoma with a relatively low number of MC1R receptors. Tumor uptake values of 64Cu-DOTA-NAPamide are found to be 4.63 +/- 0.45% and 2.49 +/- 0.31% ID/g in B16F10 and A375M xenografted melanoma at 2 h postinjection (pi), respectively. The B16F10 tumor uptake at 2 h pi is further inhibited to 2.29 +/- 0.24% ID/g, while A375M tumor uptake at 2 h pi remains 2.20 +/- 0.41% ID/g with a coinjection of excess alpha-MSH peptide. MicroPET imaging of 64Cu-DOTA-NAPamide in B16F10 tumor mice clearly shows good tumor localization. However, low A375M tumor uptake and poor tumor to normal tissue contrast were observed. This study demonstrates that 64Cu-DOTA-NAPamide is a promising molecular probe for alpha-MSH receptor positive melanoma PET imaging as well as MC1R expression imaging in living mice.

  View details for DOI 10.1021/bc060306g

  View details for Web of Science ID 000246485500021

  View details for PubMedID 17348700

• In vivo bioluminescence tumor imaging of RGD peptide-modified adenoviral vector encoding firefly luciferase reporter gene MOLECULAR IMAGING AND BIOLOGY Niu, G., Xiong, Z., Cheng, Z., Cai, W., Gambhir, S. S., Xing, L., Chen, X. 2007; 9 (3): 126-134

  Abstract

  The goal of this study is to demonstrate the feasibility of chemically modified human adenovirus (Ad) vectors for tumor retargeting.E1- and E3-deleted Ad vectors carrying firefly luciferase reporter gene under cytomegalovirus promoter (AdLuc) was surface-modified with cyclic arginine-glycine-aspartic acid (RGD) peptides through a bifunctional poly(ethyleneglycol) linker (RGD-PEG-AdLuc) for integrin alpha(v)beta(3) specific delivery. The Coxsackie and adenovirus viral receptor (CAR) and integrin alpha(v)beta(3) expression in various tumor cell lines was determined by reverse transcriptase PCR and fluorescence-activated cell sorting. Bioluminescence imaging was performed in vitro and in vivo to evaluate RGD-modified AdLuc infectivity.RGD-PEG-AdLuc abrogated the native CAR tropism and exhibited significantly enhanced transduction efficiency of integrin-positive tumors than AdLuc through intravenous administration.This approach provides a robust platform for site-specific gene delivery and noninvasive monitoring of the transgene delivery efficacy and homing.

  View details for DOI 10.1007/s11307-007-0079-2

  View details for Web of Science ID 000246175500005

  View details for PubMedID 17297551

  View details for PubMedCentralID PMC4165526

• Detection of bone metastases: Assessment of integrated FDG PET/CT imaging RADIOLOGY Taira, A. V., Herfkens, R. J., Gambhir, S. S., Quon, A. 2007; 243 (1): 204-211

  Abstract

  To retrospectively evaluate the positive predictive value (PPV) of fluorine 18 fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) in the identification of malignant bone lesions when the PET and CT findings are discordant and concordant.The study conformed to HIPAA standards, and the need for informed consent was waived by the institutional review board that approved the study. FDG PET/CT reports of 712 patients were reviewed to identify patients with malignant bone lesions. Fifty-nine patients (30 female and 29 male patients; age range, 10-82 years) with 113 lesions were analyzed. With use of confirmation from histopathologic examination or clinical follow-up, the PPVs of the integrated examination and of the stand-alone CT and PET components of the examination were calculated. The results were stratified according to cancer type, chemotherapy status, and number of bone lesions and were compared by using Fisher exact tests.Of 47 lesions with positive findings at both PET and CT, 46 were malignant and one was benign, for a PPV of 98%. Of 31 lesions with positive findings at PET and negative findings at CT, 19 were malignant and 12 were benign, for a PPV of 61%. Of 35 lesions with negative findings at PET and positive findings at CT, six were malignant and 29 were benign, for a PPV of 17%. Independently, the PPV of all lesions with positive findings at PET was significantly higher than that of all lesions with positive findings at CT. Chemotherapy status for lesions with positive findings at CT and the number of lesions per patient had a statistically significant effect on the PPV of examinations (P = .02 and P < .001, respectively).PET/CT has a very high PPV for bone metastases (98%) when the findings at PET and CT are concordant; however, in lesions with discordant PET and CT findings at the integrated examination, PPV is markedly diminished.

  View details for DOI 10.1148/radiol.2431052104

  View details for Web of Science ID 000245312500025

  View details for PubMedID 17392254

• Construction and validation of improved triple fusion reporter gene vectors for molecular imaging of living subjects CANCER RESEARCH Ray, P., Tsien, R., Gambhir, S. S. 2007; 67 (7): 3085-3093

  Abstract

  Multimodality imaging using several reporter genes and imaging technologies has become an increasingly important tool in determining the location(s), magnitude, and time variation of reporter gene expression in small animals. We have reported construction and validation of several triple fusion genes composed of a bioluminescent, a fluorescent, and a positron emission tomography (PET) reporter gene in cell culture and in living subjects. However, the bioluminescent and fluorescent components of fusion reporter proteins encoded by these vectors possess lesser activities when compared with the bioluminescent and fluorescent components of the nonfusions. In this study, we first created a mutant (mtfl) of a thermostable firefly luciferase (tfl) bearing the peroxisome localization signal to have greater cytoplasmic localization and improved access for its substrate, d-luciferin. Comparison between the three luciferases [mtfl, tfl, and firefly luciferase (fl)] both in cell culture and in living mice revealed that mtfl possessed 6- to 10-fold (in vitro) and 2-fold (in vivo) higher activity than fl. The improved version of the triple fusion vector carrying mtfl as the bioluminescent reporter component showed significantly (P < 0.05) higher bioluminescence than the previous triple fusion vectors. Of the three different red fluorescent reporter genes (jred, hcred, and mrfp1, isolated from jellyfish chromophore, coral Heteractis crispa, and coral Discosoma, respectively) evaluated, mrfp1 was able to preserve highest expression as a component of the triple fusion reporter gene for in vivo fluorescence imaging. A truncated version of wild-type herpes simplex virus 1 (HSV1) thymidine kinase gene (wttk) retained a higher expression level than the truncated mutant HSV1-sr39 TK (ttk) as the third reporter component of this improved triple fusion vector. Multimodality imaging of tumor-bearing mice using bioluminescence and microPET showed higher luciferase activity [(2.7 +/- 0.1 versus 1.9 +/- 0.1) x (10(6) p/s/cm(2)/sr)] but similar level of fluorine-18-labeled 2'-fluoro-2'-deoxyarabinofuranosyl-5-ethyluracil (18F-FEAU) uptake (1.37 +/- 0.15 versus 1.37 +/- 0.2) percentage injected dose per gram] by mtfl-mrfp1-wttk-expressing tumors compared with the fl-mrfp1-wttk-expressing tumors. Both tumors showed 4- to 5-fold higher accumulation (P < 0.05) of 18F-FEAU than fluorine-18-labeled 9-(4-fluoro-3-hydroxymethylbutyl)guanine. This improved triple fusion reporter vector will enable high sensitivity detection of lower numbers of cells from living animals using the combined bioluminescence, fluorescence, and microPET imaging techniques.

  View details for DOI 10.1158/0008-5472.CAN-06-2402

  View details for Web of Science ID 000245622900025

  View details for PubMedID 17409415

• Reproducibility of F-18-FDG microPET studies in mouse tumor xenografts JOURNAL OF NUCLEAR MEDICINE Dandekar, M., Tseng, J. R., Gambhir, S. S. 2007; 48 (4): 602-607

  Abstract

  (18)F-FDG has been used to image mouse xenograft models with small-animal PET for therapy response. However, the reproducibility of serial scans has not been determined. The purpose of this study was to determine the reproducibility of (18)F-FDG small-animal PET studies.Mouse tumor xenografts were formed with B16F10 murine melanoma cells. A 7-min small-animal PET scan was performed 1 h after a 3.7- to 7.4-MBq (18)F-FDG injection via the tail vein. A second small-animal PET scan was performed 6 h later after reinjection of (18)F-FDG. Twenty-five sets of studies were performed. Mean injected dose per gram (%ID/g) values were calculated from tumor regions of interest. The coefficient of variation (COV) from studies performed on the same day was calculated to determine the reproducibility. Activity from the second scans performed after 6 h were adjusted by subtracting the estimated residual activity from the first (18)F-FDG injection. For 7 datasets, an additional scan immediately before the second injection was performed, and residual activity from this additional delayed scan was subtracted from the activity of the second injection. COVs of both subtraction methods were compared. Blood glucose values were measured at the time of injection and used to correct the %ID/g values.The COV for the mean %ID/g between (18)F-FDG small-animal PET scans performed on the same day 6 h apart was 15.4% +/- 12.6%. The delayed scan subtraction method did not produce any significant change in the COV. Blood glucose correction increased the COV. The injected dose, tumor size, and body weight did not appear to contribute to the variability of the scans.(18)F-FDG small-animal PET mouse xenograft studies were reproducible with moderately low variability. Therefore, serial small-animal PET studies may be performed with reasonable accuracy to measure tumor response to therapy.

  View details for DOI 10.2967/jnumed.106.036608

  View details for Web of Science ID 000245647000024

  View details for PubMedID 17401098

• Combinatorial library screening for developing an improved split-firefly luciferase fragment-assisted complementation system for studying protein-protein interactions ANALYTICAL CHEMISTRY Paulmurugan, R., Gambhir, S. S. 2007; 79 (6): 2346-2353

  Abstract

  Split reporter-based bioluminescence imaging is a useful strategy for studying protein-protein as well as other intracellular interactions. We have used a combinatorial strategy to identify a novel split site for firefly luciferase with improved characteristics over previously published split sites. A combination of fragments with greater absolute signal with near-zero background signals was achieved by screening 115 different combinations. The identified fragments were further characterized by using five different interacting protein partners and an intramolecular folding strategy. Cell culture studies and imaging in living mice was performed to validate the new split sites. In addition, the signal generated by the newly identified combination of fragments (Nfluc 398/ Cfluc 394) was compared with different split luciferase fragments currently in use for studying protein-protein interactions and was shown to be markedly superior with a lower self-complementation signal and equal or higher postinteraction absolute signal. This study also identified many different combinations of nonoverlapping and overlapping firefly luciferase fragments that can be used for studying different cellular events such as subcellular localization of proteins, cell-cell fusion, and evaluating cell delivery vehicles, in addition to protein-protein interactions, both in cells and small living animals.

  View details for DOI 10.1021/ac062053q

  View details for Web of Science ID 000244867100020

  View details for PubMedID 17295448

  View details for PubMedCentralID PMC3198827

• In vivo optical bioluminescence imaging of collagen-supported cardiac cell grafts JOURNAL OF HEART AND LUNG TRANSPLANTATION Kutschka, I., Chen, I. Y., Kofidis, T., von Degenfeld, G., Sheikh, A. Y., Hendry, S. L., Hoyt, G., Pearl, J., Blau, H. M., Gambhir, S. S., Robbins, R. C. 2007; 26 (3): 273-280

  Abstract

  Histology-based survival assessment of cell grafts does not allow for in vivo follow-up. In this study we introduce two new experimental models for longitudinal in vivo survival studies of cardiac cell grafts using optical bioluminescence imaging.H9c2 cardiomyoblasts expressing both firefly luciferase (fluc) and green fluorescent protein (GFP) reporter genes were implanted into Lewis rats. In Model 1, H9c2-fluc-IRES-GFP cells (0.5 x 10(6)) were implanted into a cryoinjured abdominal wall muscle. Cells were injected using either liquid collagen (Matrigel [MG]) or phosphate-buffered saline (PBS) suspension. Cell survival was evaluated in vivo using bioluminescence imaging on days 1, 5 and 10 post-operatively. In model 2, rats underwent ligation of the left anterior descending (LAD) artery. The donor hearts were harvested, and the infarcted region was restored ex situ using 1 x 10(6) H9c2-fluc-IRES-GFP cells seeded in collagen matrix (Gelfoam [GF]) or suspended in PBS (n = 8/group). Hearts were then transplanted into the abdomen of syngeneic recipients. Optical bioluminescence imaging was performed on Days 1, 5, 8 and 14 post-operatively. After 4 weeks, immunohistologic studies were performed.For model 1, at day 5, bioluminescence signals were markedly higher for the H9c2/MG group (449 +/- 129 photons/second x 10(3)) compared with the H9c2/PBS group (137 +/- 82 photons/second x 10(3)) (p < 0.05). For model 2, bioluminescence signals were significantly (p < 0.04) higher in the H9c2/GF group compared with plain cell injection on days 5 (534 +/- 115 vs 219 +/- 34) and 8 (274 +/- 34 vs 180 +/- 23). Data were in accordance with GFP immunohistology.Optical bioluminescence is a powerful method for assessment of cardiac cell graft survival in vivo. Collagen matrices support early survival of cardiomyoblasts after transplantation into injured musculature.

  View details for Web of Science ID 000244979000010

  View details for PubMedID 17346630

• In memoriam - Tandra R. Chaudhuri, Ph.D. (1947-2006) MOLECULAR IMAGING AND BIOLOGY Gambhir, S. 2007; 9 (2): 59

  View details for DOI 10.1007/s11307-007-0081-8

  View details for Web of Science ID 000244866700001

• Standardized uptake value atlas: Characterization of physiological 2-deoxy-2-[F-18]fluoro-D-glucose uptake in normal tissues MOLECULAR IMAGING AND BIOLOGY Wang, Y., Chiu, E., Rosenberg, J., Gambhir, S. S. 2007; 9 (2): 83-90

  Abstract

  The purpose of this study was to map the distribution of 2-deoxy-2-[18F]fluoro-D-glucose (FDG) uptake in organs of patients with no known abnormalities in those tissues.We measured maximum and mean standardized uptake values (SUV) from FDG-positron emission tomography (PET)/computed tomography (CT) obtained from 98 patients (48 males and 50 females).Significant uptake (mean SUVmean>2.5) was visualized in the cerebellum (8.0+/-2.2), soft palate (2.92+/-0.86), palatine tonsils (3.45+/-1.4), lingual tonsils (3.08+/-1.05), sublingual glands (3.3+/-1.5), and testes (2.57+/-0.56). Negative correlation for FDG uptake versus age was observed for the palatine tonsils, sublingual glands, and lungs (P<0.001).Better understanding of physiological uptake throughout the body is valuable for improved interpretive accuracy and should be useful for future semi-automated comparisons to a normal SUV database.

  View details for DOI 10.1007/s11307-006-0075-y

  View details for Web of Science ID 000244866700005

  View details for PubMedID 17225983

• Impact of integrated PET/CT on variability of target volume delineation in rectal cancer TECHNOLOGY IN CANCER RESEARCH & TREATMENT Patel, D. A., Chang, S. T., Goodman, K. A., Quon, A., Thorndyke, B., Gambhir, S. S., McMillan, A., Loo, B. W., Koong, A. C. 2007; 6 (1): 31-36

  Abstract

  Several studies have demonstrated substantial variability among individual radiation oncologists in defining target volumes using computed tomography (CT). The objective of this study was to determine the impact of combined positron emission tomography and computed tomography (PET/CT) on inter-observer variability of target volume delineation in rectal cancer. We also compared the relative concordance of two PET imaging tracers, 18F-fluorodeoxyglucose (FDG) and 18F-fluorodeoxythymidine (FLT), against conventional computed tomography (CT). Six consecutive patients with locally advanced rectal cancer were enrolled onto an institutional protocol involving preoperative chemoradiotherapy and correlative studies including FDG- and FLT-PET scans acquired in the treatment position. Using these image data sets, four radiation oncologists independently delineated primary and nodal gross tumor volumes (GTVp and GTVn) for a hypothetical boost treatment. Contours were first defined based on CT alone with observers blinded to the PET images, then based on combined PET/CT. An inter-observer similarity index (SI), ranging from a value of 0 for complete disagreement to 1 for complete agreement of contoured voxels, was calculated for each set of volumes. For primary gross tumor volume (GTVp), the difference in estimated SI between CT and FDG was modest (CT SI = 0.77 vs. FDG SI = 0.81), but statistically significant (p = 0.013). The SI difference between CT and FLT for GTVp was also slight (FLT SI = 0.80) and marginally non-significant (p < 0.082). For nodal gross tumor volume, (GTVn), SI was significantly lower for CT based volumes with an estimated SI of 0.22 compared to an estimated SI of 0.70 for FDG-PET/CT (p < 0.0001) and an estimated SI of 0.70 for FLT-PET/CT (p < 0.0001). Boost target volumes in rectal cancer based on combined PET/CT results in lower inter-observer variability compared with CT alone, particularly for nodal disease. The use of FDG and FLT did not appear to be different from this perspective.

  View details for Web of Science ID 000244732600005

  View details for PubMedID 17241098

• Reporter gene imaging of protein-protein interactions in living subjects CURRENT OPINION IN BIOTECHNOLOGY Massoud, T. F., Paulmurugan, R., De, A., Ray, P., Garnbhir, S. S. 2007; 18 (1): 31-37

  Abstract

  In the past few years there has been a veritable explosion in the field of reporter gene imaging, with the aim of determining the location, duration and extent of gene expression within living subjects. An important application of this approach is the molecular imaging of interacting protein partners, which could pave the way to functional proteomics in living animals and might provide a tool for the whole-body evaluation of new pharmaceuticals targeted to modulate protein-protein interactions. Three general methods are currently available for imaging protein-protein interactions in living subjects using reporter genes: a modified mammalian two-hybrid system, a bioluminescence resonance energy transfer (BRET) system, and split reporter protein complementation and reconstitution strategies. In the future, these innovative approaches are likely to enhance our appreciation of entire biological pathway systems and their pharmacological regulation.

  View details for DOI 10.1016/j.copbio.2007.01.007

  View details for Web of Science ID 000244593000006

  View details for PubMedID 17254764

• Multimodality imaging of tumor xenografts and metastases in mice with combined small-animal PET, small-animal CT, and bioluminescence imaging JOURNAL OF NUCLEAR MEDICINE Deroose, C. M., De, A., Loening, A. M., Chow, P. L., Ray, P., Chatziioannou, A. F., Gambhir, S. S. 2007; 48 (2): 295-303

  Abstract

  Recent developments have established molecular imaging of mouse models with small-animal PET and bioluminescence imaging (BLI) as an important tool in cancer research. One of the disadvantages of these imaging modalities is the lack of anatomic information. We combined small-animal PET and BLI technology with small-animal CT to obtain fusion images with both molecular and anatomic information.We used small-animal PET/CT and BLI to detect xenografts of different cell lines and metastases of a melanoma cell line (A375M-3F) that had been transduced with a lentiviral vector containing a trimodality imaging reporter gene encoding a fusion protein with Renilla luciferase, monomeric red fluorescent protein, and a mutant herpes simplex virus type 1 thymidine kinase.Validation studies in mouse xenograft models showed a good coregistration of images from both PET and CT. Melanoma metastases were detected by 18F-FDG PET, 9-[4-(18)F-fluoro-3-(hydroxymethyl)butyl]guanine (18F-FHBG) PET, CT, and BLI and confirmed by ex vivo assays of Renilla luciferase and mutant thymidine kinase expression. 18F-FHBG PET/CT allowed detection and localization of lesions that were not seen on CT because of poor contrast resolution and were not seen on 18F-FDG PET because of higher background uptake relative to 18F-FHBG.The combination of 18F-FHBG PET, small-animal CT, and BLI allows a sensitive and improved quantification of tumor burden in mice. This technique is potentially useful for the study of the biologic determinants of metastasis and for the evaluation of novel cancer treatments.

  View details for PubMedID 17268028

• PET imaging of colorectal cancer in xenograft-bearing mice by use of an F-18-labeled T84.66 anti-carcinoembryonic antigen diabody JOURNAL OF NUCLEAR MEDICINE Cai, W., Olafsen, T., Zhang, X., Cao, Q., Gambhir, S. S., Williams, L. E., Wu, A. M., Chen, X. 2007; 48 (2): 304-310

  Abstract

  In this study, we investigated the 18F-labeled anti-carcinoembryonic antigen (CEA) T84.66 diabody, a genetically engineered noncovalent dimer of single-chain variable fragments, for small-animal PET imaging of CEA expression in xenograft-bearing mice.18F labeling of the anti-CEA T84.66 diabody (molecular mass, 55 kDa) was achieved with N-succinimidyl-4-18F-fluorobenzoate (18F-SFB). The biodistribution of the 18F-fluorobenzyl-T84.66 diabody (18F-FB-T84.66 diabody) was evaluated in athymic nude mice bearing subcutaneous LS 174T human colon carcinoma and C6 rat glioma tumors. Serial small-animal PET imaging studies were performed to further evaluate in vivo targeting efficacy and pharmacokinetics.Radiolabeling required 35 +/- 5 (mean +/- SD) min starting from 18F-SFB, and the tracer 18F-FB-T84.66 diabody was synthesized with a specific activity of 1.83 +/- 1.71 TBq/mmol. The decay-corrected radiochemical yield was 1.40% +/- 0.16% (n = 4), and the radiochemical purity was greater than 98%. The radioimmunoreactivity was 57.1% +/- 2.0%. The 18F-FB-T84.66 diabody showed rapid and high tumor uptake and fast clearance from the circulation in the LS 174T xenograft model, as evidenced by both small-animal PET imaging and biodistribution studies. High-contrast small-animal PET images were obtained as early as 1 h after injection of the 18F-FB-T84.66 diabody, and only a background level of activity accumulation was found in CEA-negative C6 tumors. The tracer exhibited predominantly renal clearance, with some activity in the liver and spleen at early time points.The 18F-labeled diabody represents a new class of tumor-specific probes for PET that are based on targeting cell surface antigen expression. The 18F-FB-T84.66 diabody can be used for high-contrast small-animal PET imaging of CEA-positive tumor xenografts. It may be translated to the clinic for PET of CEA-positive malignancies.

  View details for Web of Science ID 000244115600038

  View details for PubMedID 17268029

• 2-deoxy-2-[F-18]fluoro-D-glucose positron emission tomography/computed tomography in the management of melanoma MOLECULAR IMAGING AND BIOLOGY Iagaru, A., Quon, A., Johnson, D., Gambhir, S. S., McDougall, I. R. 2007; 9 (1): 50-57

  Abstract

  2-Deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET)/computed tomography (CT) is widely available as a powerful imaging modality, combining the ability to detect active metabolic processes and their morphologic features in a single exam. The role of FDG-PET is proven in a variety of cancers, including melanoma, but the estimates of sensitivity and specificity are based in the majority of the published studies on dedicated PET, not PET/CT. Therefore, we were prompted to review our experience with FDG-PET/CT in the management of melanoma.This is a retrospective study on 106 patients with melanoma (20-87 years old; average: 56.8 +/- 15.9), who had whole-body FDG-PET/CT at our institution from January 2003 to June 2005. Thirty-eight patients (35.9%) were women and 68 patients (64.1%) were men. Reinterpretation of the imaging studies for accuracy and data analysis from medical records were performed.All patients had the study for disease restaging. The primary tumor depth (Breslow's thickness) at initial diagnosis was available for 76 patients (71.7%) and ranged from 0.4 to 25 mm (average: 3.56 mm). The anatomic level of invasion in the skin (Clark's level) was determined for 70 patients (66%): 3, level II; 13, level III; 43, level IV; 11, level V. The administered dose of (18)F FDG ranged from 9.8 to 21.6 mCi (average: 15.4 +/- 1.8 mCi). FDG-PET/CT had a sensitivity of 89.3% [95% confidence interval (CI): 78.5-95] and a specificity of 88% (95% CI: 76.2-94.4) for melanoma detection.This study confirms the good results of FDG-PET/CT for residual/recurrent melanoma detection, as well as for distant metastases localization. PET/CT should be an integral part in evaluation of patients with high-risk melanoma, prior to selection of the most appropriate therapy.

  View details for DOI 10.1007/s11307-006-0065-0

  View details for Web of Science ID 000243545600007

  View details for PubMedID 17051322

• Non-Invasive Imaging Strategies to Visualize Gene Expression in Living Subjects Prostate Cancer – Biology, Genetics, and the New Therapeutics Iyer, M., Gambhir, S. S. edited by Chung, L., Isaacs, W., Simons, J. 2007; 2: 193–229
• Glia-Dependent TGF-Beta Signaling, Acting Independently on the TH17 Journal of Clinical Investigation Luo, J., Ho, P., Buckwater, M., Hsu, T., Lee, L., Zhang, H., Kim, D., Kim, S., Gambhir, S. S., Steinman, L., Wyss-Coray, T. 2007
• Osseus and Soft Tissue Sarcomas: When can F-18 FDG PET/CT Evaluation Provide Useful Information? European of Nuclear Medicine and Molecular Imaging Yaghoubi, S. S., Gambhir, S. S. 2007: S152
• Targeted cellular gene therapy using IL-4 secreting DCs corrects immune dysregulations and prevents diabetes in NOD mice Creusot, R. J., Yaghoubi, S., Kodama, K., Gambhir, S. S., Dang, D., Fathman, C. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2007: S17

  View details for DOI 10.1016/j.clim.2007.03.218

  View details for Web of Science ID 000247137200041

• Endothelial progenitor cells from murine embryonic stem cells: isolation and transplantation for myocardial infarction Li, Z., Wu, J., Sheikh, A., Kraft, D., Cao, F., Xie, X., Patel, M., Gambhir, S., Robbins, R., Cooke, J., Wu, J. SAGE PUBLICATIONS LTD. 2007: 152–52

  View details for Web of Science ID 000247470400044

• Differentiation and enrichment of hepatocyte-like cells from human embryonic stem cells in vitro and in vivo STEM CELLS Duan, Y., Catana, A., Meng, Y., Yamamoto, N., He, S., Gupta, S., Gambhir, S. S., Zerna, M. A. 2007; 25 (12): 3058-3068

  Abstract

  Human embryonic stem cells (hESC) may provide a cell source for functional hepatocytes. The aim of this study is to establish a viable human hepatocyte-like cell line from hESC that can be used for cell-based therapies. The differentiated hESC were enriched by transducing with a lentivirus vector containing the green fluorescent protein (GFP) gene driven by the alpha1-antitrypsin promoter; the GFP gene is expressed in committed hepatocyte progenitors and hepatocytes. GFP+ hESC were purified by laser microdissection and pressure catapulting. In addition, differentiated hESC that were transduced with a lentivirus triple-fusion vector were transplanted into NOD-SCID mice, and the luciferase-induced bioluminescence in the livers was evaluated by a charge-coupled device camera. GFP+ hESC expressed a large series of liver-specific genes, and expression levels of these genes were significantly improved by purifying GFP+ hESC; our results demonstrated that purified differentiated hESC express nearly physiological levels of liver-specific genes and have liver-specific functions that are comparable to those of primary human hepatocytes. The differentiated hESC survived and engrafted in mouse livers, and human liver-specific mRNA and protein species were detected in the transplanted mouse liver and serum at 3 weeks after transplantation. This is the first time that human albumin generated by hESC-derived hepatocytes was detected in the serum of an animal model. This also represents the first successful transplantation of differentiated hESC in an animal liver and the first bioluminescence imaging of hESC in the liver. This study is an initial step in establishing a viable hepatocyte-like cell line from hESC. Disclosure of potential conflicts of interest is found at the end of this article.

  View details for DOI 10.1634/stemcells.2007-0291

  View details for Web of Science ID 000251707200010

  View details for PubMedID 17885076

• A co-axial scanning acoustic and. photoacoustic microscope IEEE Ultrasonics Symposium Vaithilingam, S., Ma, T., Furukawa, Y., de la Zerda, A., Oralkan, O., Kamaya, A., Keren, S., Gambhir, S. S., Jeffrey, R. B., Khuri-Yakub, B. T. IEEE. 2007: 2413–2416

  View details for Web of Science ID 000254281802104

• Development of a Bicistronic Vector for Multimodality Imaging of Estrogen Receptor Activity in a Breast Cancer Model; Preliminary Application. European Journal of Nuclear Medicine and Molecular Imaging Ottobrini L, Ciana P, Moresco R Lecchi M, Belloli S, Martelli C, Todde S, Fazio F, Gambhir SS, Maggi A, Lucignani G. 2007; 35(2): 365-378
• Follicular Dendritic Sarcoma within a Focus of Castleman?s Disease. Serial FDG PET/CT in the follow up of Recurrence with Histopatholigic Confirmation. Revista Espanola Medicina Nuclear Journa Iagaru A, Mari C, Gambhir SS 2007; 26(1): 40-45
• Reporter Gene Imaging of Protein-Protein Interactions in Living Subjects. Current Opinion in Biotechnology Massoud T, Paulmurugan R, De A, Ray P, Gambhir SS 2007; 18: 31-37
• Noninvasive monitoring of ligand-dependent VEGF receptor-2 dimerization with split firefly luciferase 49th Annual Meeting of the American-Society-for-Therapeutic-Radiology-and-Oncology (ASTRO) Lee, P., Chan, C., Hua, A., Paulmurugan, R., Chan, D., Gambhir, S., Le, Q., Giaccia, A. ELSEVIER SCIENCE INC. 2007: S96–S97

  View details for Web of Science ID 000249950200173

• Integrating Non-invasive Molecular Imaging into Molecular Medicine: An Evolving Paradigm. Trends in Molecular Medicine Massoud T, Gambhir SS 2007; 13(5): 183-191
• Fusion of Gaussia Luciferase to an Engineered Anti-carcinoembryonic Antigen (CEA) Antibody for in vivo Optical Imaging. Molecular Imaging and Biology Venisnik KM, Olafsen T, Gambhir SS, Wu A 2007; 9(5): 266-277
• Studying the biodistribution of positron emission tomography reporter probes in mice NATURE PROTOCOLS Yaghoubi, S. S., Berger, F., Gambhir, S. S. 2007; 2 (7): 1752-1755

  Abstract

  Positron emission tomography (PET) reporter probes (PRPs) are used to detect PET reporter gene (PRG) expression in living subjects. This article details protocols for analyzing the biodistribution of a PRP used to detect herpes simplex virus 1 thymidine kinase (HSV1-tk) or mutant HSV1-sr39tk PRG expression. However, the methods described are generalizable to other beta- or gamma/positron-emitting probes. Accumulation of PRPs in animal tissues can be determined by counting PRP activity of isolated tissues, whereas digital whole-body autoradiography (DWBA) provides high-resolution images of PRP biodistribution in 5- to 45-microm tissue slices of killed research animals at a single time point. Biodistribution assay results may be obtained in less than a week after beginning the assay, and DWBA image acquisitions can take up to 3 months depending on the probe's radioisotope.

  View details for DOI 10.1038/nprot.2007.228

  View details for Web of Science ID 000253139200020

  View details for PubMedID 17641641

• Molecular Imaging: The Vision and Opportunity for Radiology in the Future. Radiology Hoffman J, Gambhir SS 2007; 244(1): 39-47
• Differentiation and Enrichment of Hepatocyte-Like Cells from Human Embryonic Stem Cells in Vitro and in Vivo. Stem Cells Duan Y, Catana A, Meng Y, Yamamoto N, He S, Gupta S, Gambhir SS, Zern M. 2007; 10.1634: 1-23
• Noninvasive imaging of molecular events with bioluminescent reporter genes in living subjects. Methods in molecular biology (Clifton, N.J.) Ray, P., Gambhir, S. S. 2007; 411: 131-144

  Abstract

  Bioluminescence imaging has become a very popular tool for noninvasive monitoring of fundamental biological and molecular processes in small living subjects. Luciferases are light-emitting enzymes that can generate light (known as bioluminescence) after reacting with specific substrates. The emitted light is used as a detection system for luciferase activity, which acts as a "reporter" for the activity of any regulatory elements that control its expression. These enzymes are isolated from various organisms, conveniently modified for expression in mammalian cells, and are extensively used in molecular biology and cell culture experiments. Recent advances in optical technology have opened a new dimension for in vivo application of luciferase enzymes in biomedical research. The most commonly utilized luciferases for in vivo bioluminescence are isolated from two very different sources: firefly luciferase (or beetle luciferase) and renilla luciferase (isolated from sea pansy). Although both these luciferases can produce light following interaction with the substrates, structurally and biochemically they are very different. Here we describe the methods and applications of firefly and renilla luciferases in molecular imaging using small animals.

  View details for DOI 10.1007/978-1-59745-549-7_10

  View details for PubMedID 18287643

• Drug Delivery: Keeping Tabs on Nanocarriers. Nature Nanotechnology de la Zerda, A., Gambhir, SS 2007; 2: 745-746
• F-18FDG PET/CT evaluation of osseous and soft tissue sarcomas CLINICAL NUCLEAR MEDICINE Iagaru, A., Quon, A., McDougall, T. R., Gambhir, S. S. 2006; 31 (12): 754-760

  Abstract

  Osseous and soft tissue sarcomas (OSTS) represent a histologic heterogeneous group of malignant tumors. Most of the current clinical data on the role of F-18 FDG PET in sarcomas come from patients studied with dedicated PET and less frequently with hardware fusion PET/CT. Therefore, we were prompted to review our experience with F-18 FDG PET/CT in OSTS.This is a retrospective study (January 2003-December 2005) of 44 patients with histologic diagnoses of OSTS who had F-18 FDG PET/CT at our institution. The group included 22 men and 22 women with an age range of 2 of 84 years (average, 37 +/- 20.2 years). The administered doses of F-18 FDG range 4.1 to 19.5 mCi (average, 14.3 +/- 3 mCi). Reinterpretation of the imaging studies for accuracy and data analysis from medical records was performed.The sensitivity and specificity of combined F-18 FDG PET/CT were 100% (95% confidence interval [CI] = 75.7-100) and 93.3% (95% CI = 78.7-98.1) for the primary OSTS, and 80% (95% CI = 58.4-91.9) and 86.4% (95% CI = 66.7-95.2) for metastases. When interpreted separately, CT outperformed PET for pulmonary metastases detection: CT was 76.5% sensitive and 88% specific, whereas PET was only 57.1% sensitive but 96.4% specific. For detection of other metastases, CT was 82.3% sensitive and 76% specific, with PET demonstrating 78.6% sensitivity and 92.8% specificity.Relatively similar results (except better specificity for PET and PET/CT) were noted when examining the rate of metastases detection, excluding pulmonary lesions. However, CT had a better detection rate for pulmonary metastases when compared with PET alone. A negative PET scan in the presence of suspicious CT findings in the chest cannot reliably exclude pulmonary metastases from OSTS.

  View details for Web of Science ID 000242481400004

  View details for PubMedID 17117068

• Quantitative micro positron emission tomography (PET) imaging for the in vivo determination of pancreatic islet graft survival NATURE MEDICINE Kim, S., Doudet, D. J., Studenov, A. R., Nian, C., Ruth, T. J., Gambhir, S. S., McIntosh, C. H. 2006; 12 (12): 1423-1428

  Abstract

  Islet transplantation is an attractive approach for treating type-1 diabetes, but there is a massive loss of transplanted islets. It is currently only possible to estimate islet mass indirectly, through measurement of circulating C-peptide and insulin levels. This type of estimation, however, is not sufficiently sensitive or reproducible for follow-up of individuals who have undergone islet transplantation. Here we show that islet graft survival could be assessed for 1 month in diabetic NOD mice using 9-(4-[(18)F]-fluoro-3-hydroxymethylbutyl)guanine ([(18)F]FHBG)-positron emission tomography (PET) technology, the PET signal reflecting insulin secretory capacity of transplanted islets. Expression of the gene encoding viral interleukin-10 (vIL-10), was measurable in real time with PET scanning. Additionally, we addressed the clinical potential of this approach by visualizing transplanted islets in the liver, the preferred clinical transplantation site. We conclude that quantitative in vivo PET imaging is a valid method for facilitating the development of protocols for prolonging islet survival, with the potential for tracking human transplants.

  View details for DOI 10.1038/nm1458

  View details for Web of Science ID 000242618200024

  View details for PubMedID 17143277

• Long-term monitoring of transplanted islets using positron emission tomography MOLECULAR THERAPY Lu, Y., Dang, H., Middleton, B., Campbell-Thompson, M., Atkinson, M. A., Gambhir, S. S., Tian, J., Kaufman, D. L. 2006; 14 (6): 851-856

  Abstract

  Islet transplantation can restore glucose homeostasis in those with type 1 diabetes; however, most recipients eventually lose graft function. A noninvasive method to monitor islets following transplantation would enable assessment of their survival and aid the development of therapeutics to prolong graft survival. Here, we show that recombinant lentivirus can be used to engineer human islets to express a positron emission tomography (PET) reporter gene. Following transplantation into mice, transduced islets could be imaged in vivo using microPET and a radiolabeled probe approved by the FDA for clinical use in humans. The magnitude of signal from engineered islets implanted into the axillary cavity reflected the implanted islet mass. Signals from implanted islets decreased by approximately one-half during the first few weeks following transplantation, which may reflect islet cell death shortly after transplantation. Thereafter, the magnitude of signals from the implanted islets remained fairly constant when the recipients were repetitively reimaged over 90 days. Histological analysis of the implants showed healthy islets with PET reporter-expressing cells distributed throughout the islet architecture. These studies suggest that PET imaging of lentivirus-transduced islets could provide a safe and feasible method for long-term monitoring of islet graft survival.

  View details for DOI 10.1016/j.ymthe.2006.08.007

  View details for Web of Science ID 000242723300011

  View details for PubMedID 16982215

• Proteomic analysis of reporter genes for molecular imaging of transplanted embryonic stem cells PROTEOMICS Wu, J. C., Cao, F., Dutta, S., Xie, X., Kim, E., Chungfat, N., Gambhir, S., Mathewson, S., Connolly, A. J., Brown, M., Wang, E. W. 2006; 6 (23): 6234-6249

  Abstract

  Study of stem cells may reveal promising treatment for diseases. The fate and function of transplanted stem cells remain poorly defined. Recent studies demonstrate that reporter genes can monitor real-time survival of transplanted stem cells in living subjects. We examined the effects of a novel and versatile triple fusion (TF) reporter gene construction on embryonic stem (ES) cell function by proteomic analysis. Murine ES cells were stably transduced with a self-inactivating lentiviral vector containing fluorescence (firefly luciferase; Fluc), bioluminescence (monomeric red fluorescence protein; mRFP), and positron emission tomography (herpes simplex virus type 1 truncated thymidine kinase; tTK) reporter genes. Fluorescence-activated cell sorting (FACS) analysis isolated stably transduced populations. TF reporter gene effects on cellular function were evaluated by quantitative proteomic profiling of control ES cells versus ES cells stably expressing the TF construct (ES-TF). Overall, no significant changes in protein quantity were observed. TF reporter gene expression had no effect on ES cell viability, proliferation, and differentiation capability. Molecular imaging studies tracked ES-TF cell survival and proliferation in living animals. In summary, this is the first proteomic study, demonstrating the unique potential of reporter gene imaging for tracking ES cell transplantation non-invasively, repetitively, and quantitatively.

  View details for DOI 10.1002/pmic.200600150

  View details for Web of Science ID 000242879000011

  View details for PubMedID 17080479

  View details for PubMedCentralID PMC3683542

• PET of vascular endothelial growth factor receptor expression JOURNAL OF NUCLEAR MEDICINE Cai, W., Chen, K., Mohamedali, K. A., Cao, Q., Gambhir, S. S., Rosenblum, M. G., Chen, X. 2006; 47 (12): 2048-2056

  Abstract

  For solid tumors and metastatic lesions, tumor vascularity is a critical factor in assessing response to therapy. Here we report the first example, to our knowledge, of (64)Cu-labeled vascular endothelial growth factor 121 (VEGF(121)) for PET of VEGF receptor (VEGFR) expression in vivo.VEGF(121) was conjugated with 1,4,7,10-tetraazadodecane-N,N',N'',N'''-tetraacetic acid (DOTA) and then labeled with (64)Cu for small-animal PET of mice bearing different sized U87MG human glioblastoma xenografts. Blocking experiments and ex vivo histopathology were performed to confirm the in vivo results.There were 4.3 +/- 0.2 DOTA molecules per VEGF(121), and the VEGFR2 binding affinity of DOTA-VEGF(121) was comparable to VEGF(121). (64)Cu labeling of DOTA-VEGF(121) was achieved in 90 +/- 10 min and the radiolabeling yield was 87.4% +/- 3.2%. The specific activity of (64)Cu-DOTA-VEGF(121) was 3.2 +/- 0.1 GBq/mg with a radiochemical purity of >98%. Small-animal PET revealed rapid, specific, and prominent uptake of (64)Cu-DOTA-VEGF(121) in small U87MG tumors (high VEGFR2 expression) but significantly lower and sporadic uptake in large U87MG tumors (low VEGFR2 expression). No appreciable renal clearance of (64)Cu-DOTA-VEGF(121) was observed, although the kidney uptake was relatively high likely due to VEGFR1 expression. Blocking experiments, immunofluorescence staining, and western blot confirmed the VEGFR specificity of (64)Cu-DOTA-VEGF(121).Successful demonstration of the ability of (64)Cu-DOTA-VEGF(121) to visualize VEGFR expression in vivo may allow for clinical translation of this radiopharmaceutical for imaging tumor angiogenesis and guiding antiangiogenic treatment, especially patient selection and treatment monitoring of VEGFR-targeted cancer therapy.

  View details for Web of Science ID 000242563900047

  View details for PubMedID 17138749

• Imaging mitogen-activated protein kinase function in xenograft models of prostate cancer CANCER RESEARCH Ilagan, R., Pottratz, J., Le, K., Zhang, L., Wong, S. G., Ayala, R., Iyer, M., Wu, L., Gambhir, S. S., Carey, M. 2006; 66 (22): 10778-10785

  Abstract

  Mitogen-activated protein kinases (MAPK) play important roles in malignancy. The ability to detect and quantitate MAPKs in live animal models of cancer will facilitate an understanding of disease progression. We have developed a gene expression-based imaging system that detects and quantifies MAPK activity in prostate cancer tumors implanted into severe combined immunodeficient mice. The imaging technology uses a modified version of two-step transcriptional amplification (TSTA). The tissue specificity of gene expression is imparted by an enhanced version of the prostate-specific antigen regulatory region that expresses GAL4-ELK1. GAL4-ELK1 confers MAPK specificity by activating a firefly luciferase (FLuc) reporter gene when the Ets-like transcription factor (ELK) 1 activation domain is phosphorylated by MAPK. FLuc activity in live animals was detected using the Xenogen In vivo Imaging System. We validated the TSTA-ELK1 system by analyzing its response to epidermal growth factor treatment in transfected tissue culture cells and in adenovirus (AdTSTA-ELK1)-injected prostate cancer xenograft tumors. We measured MAPK activity in two well-characterized xenograft models, CWR22 and LAPC9. Although no significant differences in MAPK levels were detected between androgen-dependent and androgen-independent xenografts, the CWR22 models display significantly higher levels of AdTSTA-ELK1 activity versus LAPC9. Western blots of tumor extracts showed that the elevated imaging signal in CWR22 xenografts correlated with elevated levels of phosphorylated extracellular signal-regulated kinase 1/2 but not p38 or c-Jun NH(2)-terminal kinase. We conclude that a gene expression-based optical imaging system can accurately detect and quantify MAPK activity in live animals.

  View details for DOI 10.1158/0008-5472.CAN-05-3577

  View details for Web of Science ID 000242264400019

  View details for PubMedID 17108114

• How molecular imaging is speeding up antiangiogenic drug development MOLECULAR CANCER THERAPEUTICS Cai, W., Rao, J., Gambhir, S. S., Chen, X. 2006; 5 (11): 2624-2633

  Abstract

  Drug development is a long process that generally spans about 10 to 15 years. The shift in recent drug discovery to novel agents against specific molecular targets highlights the need for more robust molecular imaging platforms. Using molecular probes, molecular imaging can aid in many steps of the drug development process, such as providing whole body readout in an intact system, decreasing the workload and speeding up drug development/validation, and facilitating individualized anticancer treatment monitoring and dose optimization. The main focus of this review is the recent advances in tumor angiogenesis imaging, and the targets include vascular endothelial growth factor and vascular endothelial growth factor receptor, integrin alpha(v)beta(3), matrix metalloproteinase, endoglin (CD105), and E-selectin. Through tumor angiogenesis imaging, it is expected that a robust platform for understanding the mechanisms of tumor angiogenesis and evaluating the efficacy of novel antiangiogenic therapies will be developed, which can help antiangiogenic drug development in both the preclinical stage and the clinical settings. Molecular imaging has enormous potential in improving the efficiency of the drug development process, including the specific area of antiangiogenic drugs.

  View details for DOI 10.1158/1535-7163.MCT-06-0395

  View details for Web of Science ID 000242138000004

  View details for PubMedID 17121909

• Noninvasive imaging of bone marrow stem cell homing, survival, and proliferation in. ischemic myocardium 79th Annual Scientific Session of the American-Heart-Association Lin, S., Sheikh, A. Y., Cao, F., Gambhir, S. S., Robbins, R. C., Wu, J. C. LIPPINCOTT WILLIAMS & WILKINS. 2006: 660–60

  View details for Web of Science ID 000241792804305

• Molecular imaging of cardiac stem cell mediated angiogenic gene therapy 79th Annual Scientific Session of the American-Heart-Association Xie, X., Cao, F., Sheikh, A. Y., Li, Z., Pei, X., Gambhir, S. S., Li, R., Robbins, R. C., Wu, J. C. LIPPINCOTT WILLIAMS & WILKINS. 2006: 566–66

  View details for Web of Science ID 000241792803549

• An intramolecular folding sensor for imaging estrogen receptor-ligand interactions PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Paulmurugan, R., Gambhir, S. S. 2006; 103 (43): 15883-15888

  Abstract

  Strategies for high-throughput analysis of interactions between various hormones and drugs with the estrogen receptor (ER) are crucial for accelerating the understanding of ER biology and pharmacology. Through careful analyses of the crystal structures of the human ER (hER) ligand-binding domain (hER-LBD) in complex with different ligands, we hypothesized that the hER-LBD intramolecular folding pattern could be used to distinguish ER agonists from selective ER modulators and pure antiestrogens. We therefore constructed and validated intramolecular folding sensors encoding various hER-LBD fusion proteins that could lead to split Renilla/firefly luciferase reporter complementation in the presence of the appropriate ligands. A mutant hER-LBD with low affinity for circulating estradiol was also identified for imaging in living subjects. Cells stably expressing the intramolecular folding sensors expressing wild-type and mutant hER-LBD were used for imaging ligand-induced intramolecular folding in living mice. This is the first hER-LBD intramolecular folding sensor suited for high-throughput quantitative analysis of interactions between hER with hormones and drugs using cell lysates, intact cells, and molecular imaging of small living subjects. The strategies developed can also be extended to study and image other important protein intramolecular folding systems.

  View details for DOI 10.1073/pnas.0607385103

  View details for Web of Science ID 000241568500029

  View details for PubMedID 17043219

  View details for PubMedCentralID PMC1635097

• Bifunctional antibody-Renilla luciferase fusion protein for in vivo optical detection of tumors PROTEIN ENGINEERING DESIGN & SELECTION Venisnik, K. M., Olafsen, T., Loening, A. M., Iyer, M., Gambhir, S. S., Wu, A. M. 2006; 19 (10): 453-460

  Abstract

  An anti-carcinoembryonic antigen (CEA) antibody fragment, the anti-CEA diabody, was fused to the bioluminescence enzyme Renilla luciferase (RLuc) to generate a novel optical imaging probe. Native RLuc or one of two stabilized variants (RLucC124A, RLuc8) was used as the bioluminescent moiety. A bioluminescence ELISA showed that diabody-luciferase could simultaneously bind to CEA and emit light. In vivo optical imaging of tumor-bearing mice demonstrated specific targeting of diabody-RLuc8 to CEA-positive xenografts, with a tumor:background ratio of 6.0 +/- 0.8 at 6 h after intravenous injection, compared with antigen-negative tumors at 1.0 +/- 0.1 (P = 0.05). Targeting and distribution was also evaluated by microPET imaging using (124)I-diabody-RLuc8 and confirmed that the optical signal was due to antibody-mediated localization of luciferase. Renilla luciferase, fused to biospecific sequences such as engineered antibodies, can be administered systemically to provide a novel, sensitive method for optical imaging based on expression of cell surface receptors in living organisms.

  View details for DOI 10.1093/protein/gzl030

  View details for Web of Science ID 000240544900003

  View details for PubMedID 16882674

• F-18 fluorodeoxyglucose PET/CT as an imaging tool for staging and restaging cutaneous angiosarcoma of the scalp CLINICAL NUCLEAR MEDICINE Vasanawala, M. S., Wang, Y., Quon, A., Gambhir, S. S. 2006; 31 (9): 534-537

  Abstract

  Cutaneous angiosarcoma of the scalp is a rare highly aggressive malignant tumor that typically afflicts elderly patients and commonly presents with extensive local spread and distant metastasis. Distant metastases favor lung, liver, lymph nodes, and skin. Overall, the prognosis is poor. It differs from other soft tissue sarcomas in that the size of the lesion at presentation instead of tumor grade is the important prognostic factor. Optimal treatment is yet to be determined. Wide-margin complete excision with postoperative radiotherapy has been the most effective therapy. Chemotherapy and gene therapy have been used with some success. Local extent is critical in surgical planning, especially in the head and face, and is difficult to determine accurately with clinical examination and morphologic imaging tools. We report the case of a 70-year-old man diagnosed with multifocal angiosarcoma of the scalp. PET/CT imaging with F-18 2-fluoro-2-deoxyglucose (F-18 FDG) not only showed avid FDG uptake by an angiosarcoma (SUVmax = 10.7), but also simultaneously showed local extension of multifocal lesions with periosteal involvement and excluded metastatic abdominal nodal disease. PET/CT imaging after chemotherapy and before radiation therapy showed complete resolution of FDG uptake in the scalp and osseous lesions. Evaluation of more cases of this subset of soft tissue sarcoma with FDG PET/CT may suggest a possible role in not only staging angiosarcomas to determine the extent of local as well as distant disease, but also to potentially help determine response to therapy and early recognition of local or distant recurrence.

  View details for PubMedID 16921276

• In vivo non-invasive imaging of mesenchymal stem cells after adenoviral or retroviral infection with the herpes simplex virus type-1 thymidine kinase reporter gene Roelants, V., Bertrand, L., Havaux, X., Labar, D., Bausart, R., Bol, A., Tabilio, A., Gambhir, S., Di Ianni, M., Vanoverschelde, J. SPRINGER. 2006: S163

  View details for Web of Science ID 000202967400382

• Consensus guided mutagenesis of Renilla luciferase yields enhanced stability and light output PROTEIN ENGINEERING DESIGN & SELECTION Loening, A. M., Fenn, T. D., Wu, A. M., Gambhir, S. S. 2006; 19 (9): 391-400

  Abstract

  Luciferases, which have seen expansive employment as reporter genes in biological research, could also be used in applications where the protein itself is conjugated to ligands to create probes that are appropriate for use in small animal imaging. As the bioluminescence activity of commonly used luciferases is too labile in serum to permit this application, specific mutations of Renilla luciferase, selected using a consensus sequence driven strategy, were screened for their ability to confer stability of activity in serum as well as their light output. Using this information, a total of eight favorable mutations were combined to generate a mutant Renilla luciferase (RLuc8) that, compared with the parental enzyme, is 200-fold more resistant to inactivation in murine serum and exhibits a 4-fold improvement in light output. Results of the mutational analysis were also used to generate a double mutant optimized for use as a reporter gene. The double mutant had half the resistance to inactivation in serum of the native enzyme while yielding a 5-fold improvement in light output. These variants of Renilla luciferase, which exhibit significantly improved properties compared with the native enzyme, will allow enhanced sensitivity in existing luciferase-based assays as well as enable the development of novel probes labeled with the luciferase protein.

  View details for DOI 10.1093/protein/gzl023

  View details for Web of Science ID 000240544600001

  View details for PubMedID 16857694

• F-18FDG PET/CT imaging of extramammary Paget disease of the perianal region CLINICAL NUCLEAR MEDICINE Niederkohr, R. D., Gambhir, S. S. 2006; 31 (9): 561-563

  View details for Web of Science ID 000240122400015

  View details for PubMedID 16921286

• Noninvasive imaging of islet grafts using positron-emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lu, Y., Dang, H., Middleton, B., Zhang, Z., Washburn, L., Stout, D. B., Campbell-Thompson, M., Atkinson, M. A., Phelps, M., Gambhir, S. S., Tian, J., Kaufman, D. L. 2006; 103 (30): 11294-11299

  Abstract

  Islet transplantation offers a potential therapy to restore glucose homeostasis in type 1 diabetes patients. However, islet transplantation is not routinely successful because most islet recipients gradually lose graft function. Furthermore, serological markers of islet function are insensitive to islet loss until the latter stages of islet graft rejection. A noninvasive method of monitoring islet grafts would aid in the assessment of islet graft survival and the evaluation of interventions designed to prolong graft survival. Here, we show that recombinant adenovirus can engineer isolated islets to express a positron-emission tomography (PET) reporter gene and that these islets can be repeatedly imaged by using microPET after transplantation into mice. The magnitude of signal from engineered islets implanted into the axillary cavity was directly related to the implanted islet mass. PET signals attenuated over the following weeks because of the transient nature of adenovirus-mediated gene expression. Because the liver is the preferred site for islet implantation in humans, we also tested whether islets could be imaged after transfusion into the mouse liver. Control studies revealed that both intrahepatic islet transplantation and hyperglycemia altered the biodistribution kinetics of the PET probe systemically. Although transplanted islets were dispersed throughout the liver, clear signals from the liver region of mice receiving PET reporter-expressing islets were detectable for several weeks. Viral transduction, PET reporter expression, and repeated microPET imaging had no apparent deleterious effects on islet function after implantation. These studies lay a foundation for noninvasive quantitative assessments of islet graft survival using PET.

  View details for DOI 10.1073/pnas.0603909103

  View details for Web of Science ID 000239353900035

  View details for PubMedID 16868090

  View details for PubMedCentralID PMC1544080

• Collagen matrices enhance survival of transplanted cardiomyoblasts and contribute to functional improvement of ischemic rat hearts 78th Annual Scientific Session of the American-Heart-Association Kutschka, I., Chen, I. Y., Kofidis, T., Arai, T., von Degenfeld, G., Sheikh, A. Y., Hendry, S. L., Pearl, J., Hoyt, G., Sista, R., Yang, P. C., Blau, H. M., Gambhir, S. S., Robbins, R. C. LIPPINCOTT WILLIAMS & WILKINS. 2006: I167–I173

  Abstract

  Cardiac cell transplantation is limited by poor graft viability. We aimed to enhance the survival of transplanted cardiomyoblasts using growth factor-supplemented collagen matrices.H9c2 cardiomyoblasts were lentivirally transduced to express firefly luciferase and green fluorescent protein (GFP). Lewis rats underwent ligation of the left anterior descending artery (LAD) ligation to induce an anterior wall myocardial infarction. Hearts (n=9/group) were harvested and restored ex vivo with 1 x 10(6) genetically labeled H9c2 cells either in (1) saline-suspension, or seeded onto (2) collagen-matrix (Gelfoam [GF];), (3) GF/Matrigel (GF/MG), (4) GF/MG/VEGF (10 microg/mL), or (5) GF/MG/FGF (10 microg/mL). Hearts were then abdominally transplanted into syngeneic recipients (working heart model). Controls (n=6/group) underwent infarction followed by GF implantation or saline injection. Cell survival was evaluated using optical bioluminescence on days 1, 5, 8, 14, and 28 postoperatively. At 4 weeks, fractional shortening and ejection fraction were determined using echocardiography and magnetic resonance imaging, respectively. Graft characteristics were assessed by immunohistology. Bioluminescence signals on days 5, 8, and 14 were higher for GF-based grafts compared with plain H9c2 injections (P<0.03). Signals were higher for GF/MG grafts compared with GF alone (P<0.02). GFP-positive, spindle-shaped H9c2 cells were found integrated in the infarct border zones at day 28. Left ventricular (LV) function of hearts implanted with collagen-based grafts was better compared with controls (P<0.05). Vascular endothelial growth factor or fibroblast growth factor did not further improve graft survival or heart function.Collagen matrices enhance early survival of H9c2 cardiomyoblasts after transplantation into ischemic hearts and lead to improved LV function. Further optimization of the graft design should make restoration of large myocardial infarctions by tissue engineering approaches effective.

  View details for PubMedID 16820568

• Adenoviral human BCL-2 transgene expression attenuates early donor cell death after cardiomyoblast transplantation into ischemic rat hearts 78th Annual Scientific Session of the American-Heart-Association Kutschka, I., Kofidis, T., Chen, I. Y., von Degenfeld, G., Zwierzchoniewska, M., Hoyt, G., Arai, T., Lebl, D. R., Hendry, S. L., Sheikh, A. Y., Cooke, D. T., Connolly, A., Blau, H. M., Gambhir, S. S., Robbins, R. C. LIPPINCOTT WILLIAMS & WILKINS. 2006: I174–I180

  Abstract

  Cell transplantation for myocardial repair is limited by early cell death. Gene therapy with human Bcl-2 (hBcl-2) has been shown to attenuate apoptosis in the experimental setting. Therefore, we studied the potential benefit of hBcl-2 transgene expression on the survival of cardiomyoblast grafts in ischemic rat hearts.H9c2 rat cardiomyoblasts were genetically modified to express both firefly luciferase and green fluorescent protein (mH9c2). The cells were then transduced with adenovirus carrying hBcl-2 (AdCMVhBcl-2/mH9c2). Lewis rats underwent ligation of the left anterior descending artery (LAD) to induce a sizable left ventricular (LV) infarct. Hearts were explanted and the infarcted region was restored using collagen matrix (CM) seeded with 1x10(6) mH9c2 cells (n=9) or AdCMVhBcl-2/mH9c2 cells (n=9). Control animals received CM alone (n=6) or no infarct (n=6). Restored hearts were transplanted into the abdomen of syngeneic recipients in a "working heart" model. Cell survival was evaluated using optical bioluminescence imaging on days 1, 5, 8, 14, and 28 after surgery. The left heart function was assessed 4 weeks postoperatively using echocardiography and magnetic resonance imaging. During 4 weeks after surgery, the optical imaging signal for the AdCMVhBCL2/mH9c2 group was significantly (P<0.05) higher than that of the mH9c2-control group. Both grafts led to better fractional shortening (AdCMVhBcl-2/mH9c2: 0.21+/-0.03; mH9c2: 0.21+/-0.04; control: 0.15+/-0.03; P=0.04) and ejection fraction (AdCMVhBcl-2/mH9c2: 47.0+/-6.2; mH9c2: 48.7+/-6.1; control: 34.3+/-6.0; P=0.02) compared with controls. Importantly, no malignant cells were found in postmortem histology.Transduction of mH9c2 cardiomyoblasts with AdCMVhBcl-2 increased graft survival in ischemic rat myocardium without causing malignancies. Both AdCMVhBcl-2/mH9c2 and mH9c2 grafts improved LV function.

  View details for Web of Science ID 000238688200029

  View details for PubMedID 16820569

• Evaluation of firefly luciferase biolurninescence mediated photodynarnic toxicity in cancer cells MOLECULAR IMAGING AND BIOLOGY Schipper, M. L., Patel, M. R., Gambhir, S. S. 2006; 8 (4): 218-225

  Abstract

  This work investigated whether fLuc-catalyzed oxidation of D-luciferin generates sufficient light to induce photodynamic toxicity in cancer cells.Light emission was assessed via cooled CCD (charge-coupled device) camera. Parental and fLuc expressing cancer cells were exposed to subtoxic concentrations of photosensitizers (Rose Bengal or hypericin) and D-luciferin, sunlight, or lamplight. Toxicity was assessed by MTT assay.fLuc expressing cells emitted up to 500-fold higher levels of photons than parental cell lines. Although exposure to photosensitizer and sunlight reduced survival of various cell lines, survival of fLuc expressing cells incubated with photosensitizer and D-luciferin, or photosensitizer and lamplight, did not differ significantly from parental or untreated cells.Contesting recent reports, fLuc bioluminescence does not generate sufficient photons to induce Rose Bengal or hypericin photodynamic toxicity in a range of malignant and nonmalignant cell lines, and is not suitable as a generalizable approach to antineoplastic therapy.

  View details for DOI 10.1007/s11307-006-0048-1

  View details for Web of Science ID 000239124800004

  View details for PubMedID 16791748

• Significance of one-bead-one-compound combinational chemistry NATURE CHEMICAL BIOLOGY Chen, X., Gambhir, S. S. 2006; 2 (7): 351-352

  View details for Web of Science ID 000238376400004

  View details for PubMedID 16783336

• Merkel cell carcinoma: Is there a role for 2-deoxy-2-[F-18]fluoro-D-glucose-positron emission tomography/computed tomography? MOLECULAR IMAGING AND BIOLOGY Iagaru, A., Quon, A., McDougall, I. R., Gambhir, S. S. 2006; 8 (4): 212-217

  Abstract

  2-Deoxy-2-[F-18]fluoro-D-glucose (FDG)-positron emission tomography (PET)/computed tomography (CT) is becoming widely available as a powerful imaging modality, combining the ability to detect active metabolic processes and their morphologic features in a single study. The role of FDG-PET/CT is proven in lymphoma, melanoma, colorectal carcinoma, and other cancers. However, there are rare malignancies such as Merkel cell carcinoma that can potentially be evaluated with PET/CT. We were therefore prompted to review our experience with FDG-PET/CT in the management of patients with Merkel cell carcinoma.This is a retrospective case series of six patients with Merkel cell carcinoma, 58-81 years old (average 69 +/- 8.3), who had whole-body PET/CT at our institution from January 1st, 2003 to August 31st, 2005. Two patients were women and four were men. Reinterpretation of the imaging studies for accuracy and data analysis from medical records were performed.Twelve examinations were acquired for the six patients (one patient had six PET/CT, one patient had two PET/CT, and four patients had one PET/CT). The injected FDG doses ranged 381.1-669.7 MBq (average 573.5 +/- 70.3). Four patients had the PET/CT as part of initial staging, and two patients had the exam for restaging (after surgery and XRT). A total of six Merkel lesions (pancreas, adrenal, lip, submandibular lymph nodes, cervical lymph nodes, and parapharyngeal soft tissue) were identified in three patients and confirmed on histopathological examination. The FDG uptake in these areas was intense, with maximum standardized uptake value (SUVmax) values of 5-14 (average 10.4 +/- 3.8). In one patient, the PET/CT scan identified abnormal focal distal sigmoid uptake that was biopsied and diagnosed as adenocarcinoma. Two patients had negative scans and had no clinical evidence of disease on follow-up office visits (up to one year after PET/CT).This case series suggests that FDG-PET/CT may have a promising role in the management of patients with Merkel cell carcinoma.

  View details for DOI 10.1007/s11307-006-0047-2

  View details for Web of Science ID 000239124800003

  View details for PubMedID 16724293

• "Flying through" and "flying around" a PET/CT scan: Pilot study and development of 3D integrated F-18-FDG PET/CT for virtual bronchoscopy and colonoscopy JOURNAL OF NUCLEAR MEDICINE Quon, A., Napel, S., Beaulieu, C. F., Gambhir, S. S. 2006; 47 (7): 1081-1087

  Abstract

  The objective of this pilot project was to devise a new image acquisition and processing technique to produce PET/CT images rendered in 3-dimensional (3D) volume that can then be reviewed in several 3D formats such as virtual bronchoscopy and colonoscopy "fly-throughs" and external "fly-arounds."We tested the new imaging and processing protocol on 24 patients with various malignancies to determine whether it could dependably acquire and reformat standard tomographic 2-dimensional PET/CT images into 3D renderings.This new technique added helpful information to the diagnostic interpretation for 2 of the 24 patients. Further, in the 6 patients undergoing mediastinoscopy, bronchoscopy, or endoscopy, 3D imaging helped in preprocedural planning.In this initial study, we demonstrated both the feasibility of rendering PET/CT images into 3D volumes and the potential clinical utility of this technique for diagnostic lesion characterization and preprocedural planning.

  View details for Web of Science ID 000238879300008

  View details for PubMedID 16818940

• Noninvasive evaluation of immunosuppressive drug efficacy on acute donor cell survival MOLECULAR IMAGING AND BIOLOGY Gheysens, O., Lin, S., Cao, F., Wang, D., Chen, I. Y., Rodriguez-Porcel, M., Min, J. J., Gambhir, S. S., Wu, J. C. 2006; 8 (3): 163-170

  Abstract

  The therapeutic benefits of cell transplantation may depend on the survival of sufficient numbers of grafted cells. We evaluate four potent immunosuppressive medications aimed at preventing acute donor cell death.Embryonic rat H9c2 myoblasts were stably transduced to express firefly luciferase reporter gene (H9c2-Fluc). H9c2-Fluc cells (3x10(6)) were injected into thigh muscles of Sprague-Dawley rats (N=30) treated with cyclosporine, dexamethasone, mycophenolate mofetil, tacrolimus, or saline from day -3 to day +14. Longitudinal optical bioluminescence imaging was performed over two weeks. Fluc activity was 40.0+/-12.1% (dexamethasone), 30.5+/-12.5% (tacrolimus), and 21.5+/-3.5% (mycophenolate) vs. 12.0+/-5.0% (control) and 8.3+/-5.0% (cyclosporine) at day 4 (P<0.05). However, by day 14, cell signals had decreased drastically to <10% for all groups despite drug therapy.This study demonstrates the ability of optical molecular imaging for tracking cell survival noninvasively and raises important questions with regard to the overall efficacy of immunosuppressives for prolonging transplanted cell survival.

  View details for DOI 10.1007/s11307-006-0038-3

  View details for Web of Science ID 000237754300003

  View details for PubMedID 16555032

  View details for PubMedCentralID PMC4161130

• Prostate Targeted TSTA Oncolytic Adenovirus Sato, M., Huyn, S., Powell, R., Carey, M., Gambhir, S. S., Wu, L. NATURE PUBLISHING GROUP. 2006: S117–S118

  View details for DOI 10.1016/j.ymthe.2006.08.362

  View details for Web of Science ID 000208933501231

• Preclinical safety evaluation of F-18-FHBG: A PET reporter probe for Imaging herpes simplex virus type 1 thymidine kinase (HSV1-tk) or mutant HSV1-sr39tk's expression JOURNAL OF NUCLEAR MEDICINE Yaghoubi, S. S., Couto, M. A., Chen, C. C., Polavaram, L., Cui, G. G., Sen, L. Y., Gambhir, S. S. 2006; 47 (4): 706-715

  Abstract

  9-(4-(18)F-Fluoro-3-[hydroxymethyl]butyl)guanine ((18)F-FHBG) is a sensitive and specific PET reporter probe for imaging the PET reporter genes, herpes simplex 1 thymidine kinase (HSV1-tk) and its mutant HSV1-sr39tk. (18)F-FHBG has suitable pharmacokinetics and dosimetry for clinical applications and imaging of HSV1-TK has been demonstrated in the livers of hepatocellular cancer patients.Male and female Sprague-Dawley rats and New Zealand White rabbits were divided into equal groups receiving either 14 microg/kg cold FHBG or carrier solution, for a 14-d acute toxicity assessment. We monitored body weight, food and water consumption, body temperature, cardiovascular electrical and functional indices, respiratory performance and oxygen saturation, comprehensive blood chemistry, complete blood count (CBC), and urinalysis. We conducted daily cage-side examinations for the detection of any clinical abnormalities. Tissues of the animals that were euthanized and necropsied on day 14 were prepared for histopathologic examination.No significant differences in cardiovascular and respiratory parameters, food consumption, body weight, urine components, or clinical signs attributable to test article toxicity were observed between the treatment and control groups. Any differences noted in the blood chemistry and CBC parameters were deemed to be incidental findings unrelated to the administration of the FHBG.Acute toxicity evaluation of FHBG at 100 times the expected human dose does not indicate harm to organ function or tissues. The Food and Drug Administration has approved FHBG as an Investigational New Drug.

  View details for Web of Science ID 000236593600034

  View details for PubMedID 16595506

• Real-time imaging of disruption of human heat shock protein 90/co-chaperone p23 interactions by Hsp90 inhibitors in living subjects Chan, C. T., Paulmurugan, R., Gambhir, S. S. AMER ASSOC CANCER RESEARCH. 2006

  View details for Web of Science ID 000454606204244

• Monitoring the antitumor response of naive and memory CD8 T cells in RAG1(-/-) mice by positron-emission tomography JOURNAL OF IMMUNOLOGY Su, H., Chang, D. S., Gambhir, S. S., Braun, J. 2006; 176 (7): 4459-4467

  Abstract

  Therapeutic antitumor immunity depends on a highly migratory CTL population capable of activation and trafficking between lymphoid and tumor-bearing microanatomic sites. We recently adapted positron-emission tomography gene expression imaging for noninvasive, longitudinal localization and quantitation of antitumor T lymphocyte migration in vivo. In this study, we apply this system to enumerate the temporal accumulation of naive vs memory T cells. Naive or memory OT-1 CD8(+) T cells, retrovirally marked with the sr39TK gene, were adoptively transferred into RAG1(-/-) animals bearing EL-4 or EG.7 (an OVA-expressing subline), and repetitively imaged by microPET over several weeks. Memory cells demonstrated early accumulation and apparent proliferation, with large T cell numbers at the Ag-positive tumor as early as day 1 after T cell transfer. Naive T cells did not accumulate in the E.G7 tumor until day 8, and reached only 25% of the peak levels achieved by memory T cells. Both naive and memory cells eradicated the Ag-expressing tumor at a comparable density of intratumoral T cells (2-4 x 10(6)/g). However, due to the slower rate of T cell expansion and continued tumor growth, naive cells required approximately 10-fold higher Ag-specific precursor frequency to reach a tumoricidal cell density. As recently reported, memory but not naive T cells accumulated in local lymph nodes and lungs, where they persisted as a resident population after tumor eradication. Positron-emission tomography-based immunologic imaging is a noninvasive modality providing unique and meaningful information on the dynamics of the antitumor CTL response.

  View details for Web of Science ID 000238769300073

  View details for PubMedID 16547284

• Bioluminescent-inescent monitoring of NIS-mediated I-131 ablative effects in MCF-7 Xenografts MOLECULAR IMAGING Ghosh, M., Gambhir, S. S., De, A., Nowels, K., Goris, M., Wapnir, I. 2006; 5 (2): 76-84

  Abstract

  Optical imaging has made it possible to monitor response to anticancer therapies in tumor xenografts. The concept of treating breast cancers with (131)I is predicated on the expression of the Na(+)/I- symporter (NIS) in many tumors and uptake of I- in some. The pattern of (131)I radioablative effects were investigated in an MCF-7 xenograft model dually transfected with firefly luciferase and NIS genes. On Day 16 after tumor cell implantation, 3 mCi of (131)I was injected. Bioluminescent imaging using d-luciferin and a cooled charge-coupled device camera was carried out on Days 1, 2, 3, 7, 10, 16, 22, 29, and 35. Tumor bioluminescence decreased in (131)I-treated tumors after Day 3 and reached a nadir on Day 22. Conversely, bioluminescence steadily increased in controls and was 3.85-fold higher than in treated tumors on Day 22. Bioluminescence in (131)I-treated tumors increased after Day 22, corresponding to tumor regrowth. By Day 35, treated tumors were smaller and accumulated 33% less (99m)TcO(4)(-) than untreated tumors. NIS immunoreactivity was present in <50% of (131)I-treated cells compared to 85-90% of controls. In summary, a pattern of tumor regression occurring over the first three weeks after (131)I administration was observed in NIS-expressing breast cancer xenografts.

  View details for DOI 10.2310/7290.2006.00008

  View details for PubMedID 16954021

• Peptide-labeled near-infrared quantum dots for imaging tumor vasculature in living subjects NANO LETTERS Cai, W. B., Shin, D. W., Chen, K., Gheysens, O., Cao, Q. Z., Wang, S. X., Gambhir, S. S., Chen, X. Y. 2006; 6 (4): 669-676

  Abstract

  We report the in vivo targeting and imaging of tumor vasculature using arginine-glycine-aspartic acid (RGD) peptide-labeled quantum dots (QDs). Athymic nude mice bearing subcutaneous U87MG human glioblastoma tumors were administered QD705-RGD intravenously. The tumor fluorescence intensity reached maximum at 6 h postinjection with good contrast. The results reported here open up new perspectives for integrin-targeted near-infrared optical imaging and may aid in cancer detection and management including imaging-guided surgery.

  View details for DOI 10.1021/nl052405t

  View details for Web of Science ID 000236916200015

  View details for PubMedID 16608262

• Using radiolabeled DNA as an imaging agent to recognize protein targets JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 2006; 47 (4): 557-558

  View details for Web of Science ID 000236593600014

  View details for PubMedID 16595486

• Transcriptional profiling of reporter genes used for molecular imaging of embryonic stem cell transplantation PHYSIOLOGICAL GENOMICS Wu, J. C., Spin, J. M., Cao, F., Lin, S. A., Xie, X. Y., Gheysens, O., Chen, I. Y., Sheikh, A. Y., Robbins, R. C., Tsalenko, A., Gambhir, S. S., Quertermous, T. 2006; 25 (1): 29-38

  Abstract

  Stem cell therapy offers exciting promise for treatment of ischemic heart disease. Recent advances in molecular imaging techniques now allow investigators to monitor cell fate noninvasively and repetitively. Here we examine the effects of a triple-fusion reporter gene on embryonic stem (ES) cell transcriptional profiles. Murine ES cells were stably transfected with a self-inactivating lentiviral vector carrying a triple-fusion (TF) construct consisting of fluorescence, bioluminescence, and positron emission tomography (PET) reporter genes. Fluorescence-activated cell sorting (FACS) analysis allowed isolation of stably transfected populations. Microarray studies comparing gene expression in nontransduced control ES cells vs. stably transduced ES cells expressing triple fusion (ES-TF) revealed some increases in transcriptional variability. Annotation analysis showed that ES-TF cells downregulated cell cycling, cell death, and protein and nucleic acid metabolism genes while upregulating homeostatic and anti-apoptosis genes. Despite these transcriptional changes, expression of the TF reporter gene had no significant effects on ES cell viability, proliferation, and differentiation capability. Importantly, transplantation studies in murine myocardium demonstrated the feasibility of tracking ES-TF cells in living subjects using bioluminescence and PET imaging. Taken together, this is the first study to analyze in detail the effects of reporter genes on molecular imaging of ES cells.

  View details for DOI 10.1152/physiolgenomics.00254.2005

  View details for Web of Science ID 000236722700004

  View details for PubMedID 16390873

• Self-illuminating quantum dot conjugates for in vivo imaging NATURE BIOTECHNOLOGY So, M. K., Xu, C. J., Loening, A. M., Gambhir, S. S., Rao, J. H. 2006; 24 (3): 339-343

  Abstract

  Fluorescent semiconductor quantum dots hold great potential for molecular imaging in vivo. However, the utility of existing quantum dots for in vivo imaging is limited because they require excitation from external illumination sources to fluoresce, which results in a strong autofluorescence background and a paucity of excitation light at nonsuperficial locations. Here we present quantum dot conjugates that luminesce by bioluminescence resonance energy transfer in the absence of external excitation. The conjugates are prepared by coupling carboxylate-presenting quantum dots to a mutant of the bioluminescent protein Renilla reniformis luciferase. We show that the conjugates emit long-wavelength (from red to near-infrared) bioluminescent light in cells and in animals, even in deep tissues, and are suitable for multiplexed in vivo imaging. Compared with existing quantum dots, self-illuminating quantum dot conjugates have greatly enhanced sensitivity in small animal imaging, with an in vivo signal-to-background ratio of > 10(3) for 5 pmol of conjugate.

  View details for DOI 10.1038/nbt1188

  View details for PubMedID 16501578

• In vivo visualization of embryonic stem cell survival, proliferation, and migration after cardiac delivery CIRCULATION Cao, F., Lin, S., Xie, X. Y., Ray, P., Patel, M., Zhang, X. Z., Drukker, M., Dylla, S. J., Connolly, A. J., Chen, X. Y., Weissman, I. L., Gambhir, S. S., Wu, J. C. 2006; 113 (7): 1005-1014

  Abstract

  Recent studies have shown that stem cell therapy can promote tissue regeneration; however, monitoring stem cells in vivo remains problematic owing to limitations of conventional histological assays and imaging modalities.Murine embryonic stem (ES) cells were stably transduced with a lentiviral vector carrying a novel triple-fusion (TF) reporter gene that consists of firefly luciferase, monomeric red fluorescence protein, and truncated thymidine kinase (fluc-mrfp-ttk). ES cell viability, proliferation, and differentiation ability were not adversely affected by either reporter genes or reporter probes compared with nontransduced control cells (P=NS). Afterward, 1x10(7) of ES cells carrying the TF reporter gene (ES-TF) were injected into the myocardium of adult nude rats (n=20). Control animals received nontransduced ES cells (n=6). At day 4, the bioluminescence and positron emission tomography signals in study animals were 3.7x10(7)+/-5.8x10(6) photons.s(-1).cm(-2) per steradian (sr) and 0.08+/-0.03% injected dose/g, respectively (P<0.05 versus control). Both signals increased progressively from week 1 to week 4, which indicated ES cell survival and proliferation in the host. Histological analysis demonstrated the formation of intracardiac and extracardiac teratomas. Finally, animals (n=4) that were treated with intraperitoneal injection of ganciclovir (50 mg/kg) did not develop teratomas when compared with control animals (n=4) treated with saline (1 mL/kg).This is the first study to characterize ES cells that stably express fluorescence, bioluminescence, and positron emission tomography reporter genes and monitor the kinetics of ES cell survival, proliferation, and migration. This versatile imaging platform should have broad applications for basic research and clinical studies on stem cell therapy.

  View details for DOI 10.1161/CIRCULATIONHA.105.588954

  View details for PubMedID 16476845

• Visualization of telomerase reverse transcriptase (hTERT) promoter activity using a trimodality fusion reporter construct JOURNAL OF NUCLEAR MEDICINE Padmanabhan, P., Otero, J., Ray, P., Paulmurugan, R., Hoffman, A. R., Gambhir, S. S., Biswal, S., Ulaner, G. A. 2006; 47 (2): 270-277

  Abstract

  Our goal was to noninvasively measure chemotherapy-induced changes in the expression of critical tumor growth genes. To achieve this goal, we used radionuclide and optical methods to measure changes in human telomerase reverse transcriptase (hTERT) gene expression in tumor cells before and after 5-fluorouracil treatment.A fusion reporter construct, containing humanized Renilla luciferase (hrl, for bioluminescent imaging), monomeric red fluorescence protein 1 (mrfp1, for fluorescent imaging), and a truncated thymidine kinase (ttk, for imaging of radiolabeled acycloguanosines), was placed under the control of hTERT promoter fragments. These constructs were introduced into tumor cell lines with and without hTERT expression. Transfected cells were treated with 5-fluorouracil, a chemotherapeutic that decreases hTERT gene expression, and treatment-induced changes in hTERT promoter activity were imaged.When the fusion construct is introduced into cell lines that express hTERT, all 3 reporter systems are highly expressed and hTERT promoter activity can be visualized. Cell lines lacking hTERT transcription show no significant reporter expression. Decreases in hTERT gene expression caused by 5-fluorouracil treatment could be visualized in living 293T cells by both fluorescent microscopy and bioluminescent imaging.hTERT promoter activity can be monitored by 1 radionuclide and 2 optical reporter systems using a single reporter construct. This in vitro study provides evidence that our multimodality reporter construct can be used to study the expression of a critical tumor growth gene in living subjects.

  View details for Web of Science ID 000235283500027

  View details for PubMedID 16455633

• Diagnosis of aseptic deep venous thrombosis of the upper extremity in a cancer patient using fluorine-18 fluorodeoxyglucose positron emission tomography/computerized tomography (FDG PET/CT) ANNALS OF NUCLEAR MEDICINE Do, B., Mari, C., Biswal, S., Kalinyak, J., Quon, A., Gambhir, S. S. 2006; 20 (2): 151-155

  Abstract

  We describe a patient with a history of recurrent squamous cell carcinoma of the tongue and abnormal FDG uptake in the left arm during a re-staging FDG PET/CT. After revision of the patient's clinical history, tests and physical exam, the abnormal FDG uptake was found to correspond to an extensive aseptic deep venous thrombosis of the upper extremity.

  View details for Web of Science ID 000236242700010

  View details for PubMedID 16615425

• Noninvasive indirect imaging of vascular endothelial growth factor gene expression using bioluminescence imaging in living transgenic mice PHYSIOLOGICAL GENOMICS Wang, Y. L., Iyer, M., Annala, A., Wu, L., Carey, M., Gambhir, S. S. 2006; 24 (2): 173-180

  Abstract

  Vascular endothelial growth factor (VEGF) plays a critical role in the early activation of stromal tissues during wound healing and tumor growth. We report the use of a two-step transcriptional amplification (TSTA) approach to augment the transcriptional activity of the relatively weak VEGF promoter (pVEGF) using firefly luciferase (fl) reporter gene and bioluminescence imaging (BLI). In cell culture, we demonstrate that TSTA-based fl gene expression can be significantly enhanced over the direct one-step system. Using a transgenic mouse model (pVEGF-TSTA-fl), we demonstrate the induction of VEGF gene expression using a wound-healing model and a subcutaneous mammary tumor model. In skin-wounding experiments, pVEGF-induced fl expression in the wound lesion is detected on days 4 and 5 and peaks on days 15-22. Furthermore, the bioluminescence signal shows good correlation with the endogenous VEGF protein levels in the wound tissue (r2 = 0.70). In the mammary tumor model, fl expression is detected on day 3, peaks at day 17, and declines thereafter. These results support the use of noninvasive BLI for the longitudinal monitoring of VEGF induction during wound healing and tumor progression, and this mouse model should find use in various applications in which it is important to noninvasively study VEGF gene expression.

  View details for DOI 10.1152/physiolgenomics.00308.2004

  View details for Web of Science ID 000234590300011

  View details for PubMedID 16410544

• Quantitative PET imaging of tumor integrin alpha(v)beta(3) expression with F-18-FRGD2 JOURNAL OF NUCLEAR MEDICINE Zhang, X. Z., Xiong, Z. M., Wu, Y., Cai, W. B., Tseng, J. R., Gambhir, S. S., Chen, X. Y. 2006; 47 (1): 113-121

  Abstract

  The development of noninvasive methods to visualize and quantify integrin alpha(v)beta(3) expression in vivo appears to be crucial for the success of antiangiogenic therapy based on integrin antagonism. Precise documentation of integrin receptor levels will allow appropriate selection of patients who will most likely benefit from an antiintegrin treatment regimen. Imaging can also be used to provide an optimal dosage and time course for treatment based on receptor occupancy studies. In addition, imaging integrin expression will be important to evaluate antiintegrin treatment efficacy and to develop new therapeutic drugs with favorable tumor targeting and in vivo kinetics. We labeled the dimeric RGD peptide E[c(RGDyK)](2) with (18)F and evaluated its tumor-targeting efficacy and pharmacokinetics of (18)F-FB-E[c(RGDyK)](2) ((18)F-FRGD2).E[c(RGDyK)](2) was labeled with (18)F by conjugation coupling with N-succinimidyl-4-(18)F-fluorobenzoate ((18)F-SFB) under a slightly basic condition. The in vivo metabolic stability of (18)F-FRGD2 was determined. The diagnostic value after injection of (18)F-FRGD2 was evaluated in various xenograft models by dynamic microPET followed by ex vivo quantification of tumor integrin level.Starting with (18)F(-) Kryptofix 2.2.2./K(2)CO(3) solution, the total reaction time for (18)F-FRGD2, including final high-performance liquid chromatography purification, is about 200 +/- 20 min. Typical decay-corrected radiochemical yield is 23% +/- 2% (n = 20). (18)F-FRGD2 is metabolically stable. The binding potential extrapolated from graphical analysis of PET data and Logan plot correlates well with the receptor density measured by sodium dodecyl sulfate polyacrylamide electrophoresis and autoradiography in various xenograft models. The tumor-to-background ratio at 1 h after injection of (18)F-FRGD2 also gives a good linear relationship with the tumor tissue integrin level.The dimeric RGD peptide tracer (18)F-FRGD2, with high integrin specificity and favorable excretion profile, may be translated into the clinic for imaging integrin alpha(v)beta(3) expression. The binding potential calculated from simplified tracer kinetic modeling such as the Logan plot appears to be an excellent indicator of tumor integrin density.

  View details for Web of Science ID 000234679300022

  View details for PubMedID 16391195

• Bioluminescence Reporter Gene Imaging in Small Animal Models of Cancer New Techniques in Oncological Imaging Massoud, T. F., Gambhir, S. S. edited by Padhani, A., Choyke, P. Taylor and Francis Books. 2006: 291–317
• Imaging Gene Expression: Concepts and Future Outlook Diagnostic Nuclear Medicine Gambhir, S. S., Schipper, M. edited by Schiepers, C. Springer Verlag: Berlin Heidelberg New York. 2006; 2: 313–342
• Near-Infrared Fluorescent Deoxyglucose Analogue for Tumor Optical Imaging in Cell Culture and Living Mice Bioconuj Chem Cheng Z, Levi J, Xiong Z, Gheysens O, Keren S, Chen X, Gambhir SS 2006; 17: ASAP
• Non-Invasive Imaging of Islet Grafts Using Positron Emission Tomography Proceedings of the National Academy of Sciences (USA) Lu, Y., Dang, H., Middleton, B., Zhang, Z., Washburn, L., Stout, D. B., Campbell-Thompson, M., Atkinson, M. A., Phelps, M., Gambhir, S. S., Tian, J., Kaufman, D. L. 2006: 11294-11299
• A hybrid microCT scanner for image-guided conformal radiotherapy of small animals Graves, E. E., Chatterjee, R., Gambhir, S. S., Contag, C. H., Boyer, A. L. ELSEVIER SCIENCE INC. 2006: S707–S708

  View details for DOI 10.1016/j.ijrobp.2006.07.1299

  View details for Web of Science ID 000241221602391

• Creating self-illuminating quantum dot conjugates Creating self-illuminating quantum dot conjugates So, M., et al 2006; 3: 1160-1164
• Bioluminescence imaging of systemic tumor targeting using a prostate-specific lentiviral vector HUMAN GENE THERAPY Iyer, M., Salazar, F. B., Wu, L., Carey, M., Gambhir, S. S. 2006; 17 (1): 125-132

  Abstract

  Developments in vector design using tissue-specific and tumor-specific promoters have led to significant improvements in tumor-targeting strategies. These developments combined with the ability to monitor gene expression by molecular imaging have facilitated the detection and prolonged monitoring of disease progression in small-animal models. Bioluminescence imaging offers a convenient and sensitive platform for monitoring gene expression patterns in preclinical models of gene therapy. Targeting a specific subset of cells/tissues via systemic delivery of vectors would be highly beneficial in gene therapy protocols. Using a two-step transcriptional amplification (TSTA)-based lentiviral vector (LV-TSTA), we demonstrate specific targeting of prostate tumors in vivo after systemic administration of lentivirus. Four days after intravenous administration of LV-TSTA into adult severe combined immunodeficient (SCID) mice (n=5) carrying subcutaneous prostate tumors, we found significant levels of transduction at the tumor site when compared with other organs (p<0.05). Gene expression was sustained in the tumor for up to 3 weeks (7.3x10(4)+/-2x10(4) photons/ sec/cm2/steradian (p/sec/cm2/sr) on day 4 and 7.0x10(4)+/-4x10(4) p/sec/cm2/sr on day 21). Low levels of transduction were also observed in the spleen and liver (5.0x10(2)+/-1.7x10(2) p/sec/cm2/sr). The results from this study support the use of TSTA-based lentiviral vectors for prostate tumor targeting after systemic delivery. Noninvasive imaging using such vectors should be useful for monitoring long-term gene expression in gene therapy applications.

  View details for Web of Science ID 000234720400012

  View details for PubMedID 16409131

• Long-Term Monitoring of Transplanted Islets using Positron Emission Tomography. Molecular Therapy Lu Y, Dang H, Middleton B, Thompson M, Atkinson M, Gambhir SS, Tian J, Kaufman D. 2006; 14: 851-856
• Visualization of Telomerase Reverse Transcriptase (hTERT) Promoter Activity Using a Tri-modality Fusion Reporter Construct Journal of Nuclear Medicine Padmanabhan P, Ray P, Paulmurugan R, Otero J, Hoffman AR, Gambhir SS, Biswal S, Ulaner GA 2006; 47: 270-277
• Imaging Mitogen-Activated Protein Kinase Function in Xenograft Models of Prostate Cancer. Cancer Research Ilagan R, Pottratz J, Le K, Zhang L, Wong SG, Ayala R, Iyer M, Gambhir SS, Carey M. 2006; 66: 10778-10785
• HaloTag protein-mediated site-specific conjugation of bioluminescent proteins to quantum dots ANGEWANDTE CHEMIE-INTERNATIONAL EDITION Zhang, Y., So, M., Loening, A. M., Yao, H., Gambhir, S. S., Rao, J. 2006; 45 (30): 4936-4940

  View details for DOI 10.1002/anie.200601197

  View details for PubMedID 16807952

• Non-invasive Indirect Imaging of VEGF Gene Expression using Bioluminescence Imaging in Living Transgenic Mice. Physiological Genomics Wang Y, Iyer M, Annala A, Wu L, Carey M, Gambhir SS. 2006; 24: 173-180
• Measuring herpes simplex virus thymidine kinase reporter gene expression in vitro NATURE PROTOCOLS Yaghoubi, S. S., Gambhir, S. S. 2006; 1 (4): 2137-2142

  Abstract

  The herpes simplex 1 virus thymidine kinase (HSV1-tk) positron emission tomography (PET) reporter gene (PRG) or its mutant HSV1-sr39tk are used to investigate intracellular molecular events in cultured cells and for imaging intracellular molecular events and cell trafficking in living subjects. Two in vitro methods are available to assay gene expression of HSV1-tk or HSV1-sr39tk in cells or tissues. One method determines the level of HSV1-TK or HSV1-sr39TK enzyme activity in cell or tissue lysates by measuring the amount of the radiolabeled substrates that have been phosphorylated by these enzymes in a fixed amount of cell lysate protein after a fixed incubation time. The other method, called the 'cell-uptake assay', takes into account the natural uptake and efflux characteristics of the radiolabeled substrate by specific cells, in addition to the level of HSV1-TK or HSV1-sr39TK activity. Both of these assays can be used to validate molecular models in cultured cells, prior to studying them in living research subjects. Each of these assays can be completed in one day.

  View details for DOI 10.1038/nprot.2006.334

  View details for Web of Science ID 000251155500058

  View details for PubMedID 17487205

• Creating self-illuminating quantum dot conjugates NATURE PROTOCOLS So, M., Loening, A. M., Gambhir, S. S., Rao, J. 2006; 1 (3): 1160-1164

  Abstract

  Semiconductor quantum dots are inorganic fluorescent nanocrystals that, because of their unique optical properties compared with those of organic fluorophores, have become popular as fluorescent imaging probes. Although external light excitation is typically required for imaging with quantum dots, a new type of quantum dot conjugate has been reported that can luminesce with no need for external excitation. These self-illuminating quantum dot conjugates can be prepared by coupling of commercially available carboxylate-presenting quantum dots to the light-emitting protein Renilla luciferase. When the conjugates are exposed to the luciferase's substrate coelenterazine, the energy released by substrate catabolism is transferred to the quantum dots through bioluminescence resonance energy transfer, leading to quantum dot light emission. This protocol describes step-by-step procedures for the preparation and characterization of these self-illuminating quantum dot conjugates. The preparation process is relatively simple and can be done in less than 2 hours. The availability of self-illuminating quantum dot conjugates will provide many new possibilities for in vivo imaging and detection, such as monitoring of in vivo cell trafficking, multiplex bioluminescence imaging and new quantum dot-based biosensors.

  View details for DOI 10.1038/nprot.2006.162

  View details for PubMedID 17406398

• Quantitative Micro Positron Emission Tomography (PET) Imaging for the in vivo Determination of Pancreatic Islet Graft Survival. Nature Medicine Kim SJ, Doudet DJ, Studenov AR, Nian C, ruth TJ, Gambhir SS, McIntosh CH 2006; 12: 1423-1428
• Self-Illuminating Quantum Dot Conjugates for in vivo Imaging. Nature Biotechnology So M, Xu C, Loening A, Gambhir SS, Rao J 2006; 24(3): 339-343
• Bifunctional Antibody-Renilla Luciferase Fusion Protein for in Vivo Optical Detection of Tumors. Protein Engineering, Design & Selection Venisnik KM, Olafsen T, Loening AM, Iyer M, Gambhir SS, Wu AM 2006; 19: 453-460
• Consensus Guided Mutagenesis of Renilla Luciferase Yields Enhanced Stability and Light Output. Protein Engineering, Design & Selection Loening AM, Fenn TD, Wu, AM, Gambhir SS 2006; 19: 391-400
• Monitoring the Anti-tumor Response of Naive and Memory CD8 T Cells in RAG1-/-Mice by Positron-Emission Tomography. The Journal of Immunology Su H, Chang DS, Gambhir SS, Braun J 2006; 176(7): 4459-4467
• Non-invasive Imaging of Islet Grafts using Positron Emission Tomography. Proceedings of the National Academy of Sciences (USA) Lu Y, Dang H, Middleton B, Zhang Z, Washburn L, Stout DB, Campbell-Thompson M, Atkinson MA, Phelps M, Gambhir SS, Tian J, Kaufman DL 2006; 103: 11294-11299
• PET imaging of herpes simplex virus type 1 thymidine kinase (HSV1-tk) or mutant HSV1-sr39tk reporter gene expression in mice and humans using [F-18] FHBG NATURE PROTOCOLS Yaghoubi, S. S., Gambhir, S. S. 2006; 1 (6): 3069-3075

  Abstract

  The herpes simplex virus type 1 thymidine kinase (HSV1-tk) positron emission tomography (PET) reporter gene (PRG) or its mutant HSV1-sr39tk are used to investigate intracellular molecular events in cultured cells and to image intracellular molecular events and cell trafficking in living subjects. The expression of these PRGs can be imaged using 18F- or 124I-radiolabeled acycloguanosine or pyrimidine analog PET reporter probes (PRPs). This protocol describes the procedures for imaging HSV1-tk or HSV1-sr39tk PRG expression in living subjects with the acycloguanosine analog 9-4-[18F]fluoro-3-(hydroxymethyl)butyl]guanine ([18F]FHBG). [18F]FHBG is a high-affinity substrate for the HSV1-sr39TK enzyme with relatively low affinity for mammalian TK enzymes, resulting in improved detection sensitivity. Furthermore, [18F]FHBG is approved by the US Food and Drug Administration as an investigational new imaging agent and has been shown to detect HSV1-tk transgene expression in the liver tumors of patients. MicroPET imaging of each small animal can be completed in approximately 1.5 h, and each patient imaging session takes approximately 3 h.

  View details for DOI 10.1038/nprot.2006.459

  View details for Web of Science ID 000251155700069

  View details for PubMedID 17406570

• Effects of epigenetic modulation on reporter gene expression: implications for stem cell imaging. FASEB journal Krishnan, M., Park, J. M., Cao, F., Wang, D., Paulmurugan, R., Tseng, J. R., Gonzalgo, M. L., Gambhir, S. S., Wu, J. C. 2006; 20 (1): 106-108

  Abstract

  Tracking stem cell localization, survival, differentiation, and proliferation after transplantation in living subjects is essential for understanding stem cell biology and physiology. In this study, we investigated the long-term stability of reporter gene expression in an embryonic rat cardiomyoblast cell line and the role of epigenetic modulation on reversing reporter gene silencing. Cells were stably transfected with plasmids carrying cytomegalovirus promoter driving firefly luciferase reporter gene (CMV-Fluc) and passaged repeatedly for 3-8 months. Within the highest expressor clone, the firefly luciferase activity decreased progressively from passage 1 (843+/-28) to passage 20 (250+/-10) to passage 40 (44+/-3) to passage 60 (3+/-1 RLU/microg; P<0.05 vs. passage 1). Firefly luciferase activity was maximally rescued by treatment with 5-azacytidine (DNA methyltransferase inhibitor) compared with trichostatin A (histone deacetylase inhibitor) and retinoic acid (transcriptional activator; P<0.05). Increasing dosages of 5-azacytidine treatment led to higher levels of firefly luciferase mRNA (RT-PCR) and protein (Western blots) and inversely lower levels of methylation in the CMV promoter (DNA nucleotide sequence). These in vitro results were extended to in vivo bioluminescence imaging (BLI) of cell transplant in living animals. Cells treated with 5-azacytidine were monitored for 2 wk compared with 1 wk for untreated cells (P<0.05). These findings should have important implications for reporter gene-based imaging of stem cell transplantation.

  View details for PubMedID 16246867

• Imaging chemically modified adenovirus for targeting tumors expressing integrin alpha(v)beta(3) in living mice with mutant herpes simplex virus type 1 thymidine kinase PET reporter gene JOURNAL OF NUCLEAR MEDICINE Xiong, Z. M., Cheng, Z., Zhang, X. Z., Patel, M., Wu, J. C., Gambhir, S. S., Chen, X. Y. 2006; 47 (1): 130-139

  Abstract

  The aim of this study was to change adenovirus tropism by chemical modification of the fiber knobs with PEGylated RGD peptide for targeting integrin alpha(v)beta(3) that is uniquely or highly expressed in tumor cells and neovasculature of tumors of various origins.The first generation Ad (Ad) vector, which expresses the herpes simplex virus type 1 mutant thymidine kinase (HSV1-sr39tk) gene under the control of cytomegalovirus (CMV) promoter was conjugated with poly(ethylene glycol) (PEG) or RGD-PEG. The transduction efficiency of Ads (Adtk, PEG-Adtk, and RGD-PEG-Adtk) into different types of cells (293T, MCF7, MDA-MB-435, and U87MG) was analyzed and quantified by thymidine kinase (TK) assay using 8-(3)H-penciclovir (8-(3)H-PCV) as substrate. The in vivo infectivity of the Ad vectors after intravenous administration into integrin alpha(v)beta(3)-positive U87MG and MDA-MB-435 tumor-bearing athymic nude mice was measured by both noninvasive microPET using 9-[4-(18)F-fluoro-3-(hydroxymethyl)butyl]guanine ((18)F-FHBG) as a reporter probe and ex vivo TK assay of the tumor and tissue homogenates.PEGylation completely abrogated coxsackievirus and adenovirus receptor (CAR)-knob interaction and the infectivity of PEG-Adtk is significantly lower than that of unmodified Adtk in CAR-positive cells. RGD-PEG-modified virus (RGD-PEG-Adtk) had significantly higher infectivity than PEG-Adtk and the extent of increase is related to both CAR and integrin alpha(v)beta(3) expression levels. (18)F-FHBG had minimal nonspecific uptake in the liver and tumors that are void of sr39tk. Mice preinjected intravenously with unmodified Adtk resulted in high hepatic uptake and moderate tumor accumulation of the tracer. In contrast, RGD-PEG-Adtk administration resulted in significantly lower liver uptake without compromising the tumor accumulation of (18)F-FHBG. Expression of TK in the liver and tumor homogenates corroborated with the magnitude of (18)F-FHBG uptake quantified by noninvasive microPET. Analysis of liver and tumor tissue integrin level confirmed that RGD-integrin interaction is responsible for the enhanced tumor infectivity of RGD-PEG-Adtk.The results of this study suggest that RGD-PEG conjugation is an effective way to modify Ad vector tropism for improved systemic gene delivery. Noninvasive PET and (18)F-FHBG are able to monitor in vivo transfectivity of both Adtk and RGD-PEG-Adtk vectors in the liver and tumors after intravenous injection.

  View details for PubMedID 16391197

• Image-guided cardiac cell delivery using high-resolution small-animal ultrasound MOLECULAR THERAPY Rodriguez-Porcel, M., Gheysens, O., Chen, I. Y., Wu, J. C., Gambhir, S. S. 2005; 12 (6): 1142-1147

  Abstract

  Open-chest cardiac injection is the preferred delivery method for cardiac gene and stem cell therapy in small animals, but it is invasive and the operator is unable to see the actual delivery site. High-resolution ultrasound has recently been developed for small-animal imaging. We tested the hypothesis that image-guided cardiac cell delivery using high-resolution ultrasound guidance is feasible and reproducible. Sprague-Dawley rats (n = 11) were imaged using high-resolution ultrasound, and stably transfected cardiomyoblasts (plasmid-CMV-firefly luciferase) were injected into the anterior cardiac wall under ultrasound guidance (parasternal long-axis view), using a 28-gauge needle. After injection, bioluminescence imaging was performed using a cooled charged-coupled camera. Injection was successful in all animals and was associated with no mortality. The signal detected was positively correlated with the amount of cells transplanted (R(2) = 0.94, P = 0.03) and highly correlated with ex vivo assays (R(2) = 0.82). In addition, the optical signal could be followed longitudinally using bioluminescence imaging. Ultrasound image-guided cardiac cell delivery is an effective, safe, and reproducible way to perform cell delivery to a specific myocardial region and can be combined with assessment of cardiac function. We are confident that the use of these technologies will play a significant role in the future of gene and cell therapy.

  View details for DOI 10.1016/j.ymthe.2005.07.532

  View details for Web of Science ID 000233864700017

  View details for PubMedID 16111921

• Multimodality tumor imaging targeting integrin alphavbeta3. BioTechniques Cai, W., Sam Gambhir, S., Chen, X. 2005; 39 (6): S14-25

  Abstract

  The cell adhesion molecule integrin alphavbeta3 is an important player in the process of tumor angiogenesis and metastasis. Antibodies, peptides, peptidomimetics, and small molecule antagonists against integrin alphavbeta3 have been shown to induce endothelial apoptosis, to inhibit tumor angiogenesis, and to increase endothelial permeability. The ability to quantitatively image integrin alphavbeta3 expression in vivo in a noninvasive manner may shed new light into the mechanism of angiogenesis and antiangiogenic treatment efficacy based on integrin antagonism. Tumor integrin expression imaging will also aid in lesion detection, patient stratification, new anti-integrin drug development/validation, as well as treatment monitoring and optimization. This review summarizes the recent advances in multimodality imaging of tumor integrin alphavbeta3 expression using magnetic resonance imaging (MRI), ultrasound, near-infrared (NIR) fluorescence, single photon emission computed tomography (SPECT), and positron emission tomography (PET).

  View details for DOI 10.2144/000112091

  View details for PubMedID 20158499

• Clinical molecular imaging and therapy - moving ahead together EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Bengel, F. M., Gambhir, S. S. 2005; 32: S323-S323

  View details for DOI 10.1007/s00259-005-1959-9

  View details for Web of Science ID 000203011200001

  View details for PubMedID 16258761

• Applications of molecular imaging in cancer gene therapy CURRENT GENE THERAPY Iyer, M., Sato, M., Johnson, M., Gambhir, S. S., Wu, L. 2005; 5 (6): 607-618

  Abstract

  Gene-based therapy is a promising and flexible therapeutic approach to manage diverse types of cancer. The lack of convincing therapeutic success of current gene therapy protocols in part, can be attributed to the inability to monitor gene expression at the targeted site in the living subject. Linking molecular imaging to gene therapy will enable real-time assessment of the therapeutic process and the refinement of treatment protocols. This review will cover two common imaging modalities, positron emission tomography (PET) and bioluminescence imaging (BLI), used in pre-clinical and clinical gene therapy applications. Strategies to develop more specific and robust cancer gene therapy and imaging approaches will be discussed. Coupling PET to gene therapy of cancer has already been implemented in several clinical studies. This approach would help to improve the efficacy and safety of future gene therapy clinical trials.

  View details for Web of Science ID 000233920100007

  View details for PubMedID 16457650

• Gene therapy imaging in patients for oncological applications EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Penuelas, I., Haberkorn, U., Yaghoubi, S., Gambhir, S. S. 2005; 32: S384-S403

  Abstract

  Thus far, traditional methods for evaluating gene transfer and expression have been shown to be of limited value in the clinical arena. Consequently there is a real need to develop new methods that could be repeatedly and safely performed in patients for such purposes. Molecular imaging techniques for gene expression monitoring have been developed and successfully used in animal models, but their sensitivity and reproducibility need to be tested and validated in human studies. In this review, we present the current status of gene therapy-based anticancer strategies and show how molecular imaging, and more specifically radionuclide-based approaches, can be used in gene therapy procedures for oncological applications in humans. The basis of gene expression imaging is described and specific uses of these non-invasive procedures for gene therapy monitoring illustrated. Molecular imaging of transgene expression in humans and evaluation of response to gene-based therapeutic procedures are considered. The advantages of molecular imaging for whole-body monitoring of transgene expression as a way to permit measurement of important parameters in both target and non-target organs are also analyzed. The relevance of this technology for evaluation of the necessary vector dose and how it can be used to improve vector design are also examined. Finally, the advantages of designing a gene therapy-based clinical trial with imaging fully integrated from the very beginning are discussed and future perspectives for the development of these applications outlined.

  View details for DOI 10.1007/s00259-005-1928-3

  View details for Web of Science ID 000203011200005

  View details for PubMedID 16180032

• Visualization of a primary anti-tumor immune response by positron emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Shu, C. Y., Guo, S. L., Kim, Y. J., Shelly, S. M., Nijagal, A., Ray, P., Gambhir, S. S., Radu, C. G., Witte, O. N. 2005; 102 (48): 17412-17417

  Abstract

  Current methodologies that monitor immune responses rely on invasive techniques that sample tissues at a given point in time. New technologies are needed to elucidate the temporal patterns of immune responses and the spatial distribution of immune cells on a whole-body scale. We describe a noninvasive, quantitative, and tomographic approach to visualize a primary anti-tumor immune response by using positron emission tomography (PET). Bone marrow chimeric mice were generated by engraftment of hematopoietic stem and progenitor cells transduced with a trifusion reporter gene encoding synthetic Renilla luciferase (hRluc), EGFP, and Herpes virus thymidine kinase (sr39TK). Mice were challenged with the Moloney murine sarcoma and leukemia virus complex (M-MSV/M-MuLV), and the induced immune response was monitored by using PET. Hematopoietic cells were visualized by using 9-[4-[(18)F]fluoro-3-(hydroxymethyl)butyl]guanine ([(18)F]FHBG), a radioactive substrate specific for the sr39TK PET reporter protein. Immune cell localization and expansion were seen at the tumor and draining lymph nodes (DLNs). 2-[(18)F]fluoro-2-deoxy-D-glucose ([(18)F]FDG), which is sequestered in metabolically active cells, was used to follow tumor growth and regression. Elevated glucose metabolism was also seen in activated lymphocytes in the DLNs by using the [(18)F]FDG probe. When M-MSV/M-MuLV-challenged mice were treated with the immunosuppressive drug dexamethasone, activation and expansion of immune cell populations in the DLNs could no longer be detected with PET imaging. The method we describe can be used to kinetically measure the induction and therapeutic modulations of cell-mediated immune responses.

  View details for DOI 10.1073/pnas.0508698102

  View details for Web of Science ID 000233762000028

  View details for PubMedID 16293690

  View details for PubMedCentralID PMC1283986

• Reproducibility of 3 '-deoxy-3 '-F-18-fluorothymidine MicroPET studies in tumor xenografts in mice JOURNAL OF NUCLEAR MEDICINE Tseng, J. R., Dandekar, M., Subbarayan, M., Cheng, Z., Park, J. M., Louie, S., Gambhir, S. S. 2005; 46 (11): 1851-1857

  Abstract

  3'-Deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) has been used to image tumor proliferation in preclinical and clinical studies. Serial microPET studies may be useful for monitoring therapy response or for drug screening; however, the reproducibility of serial scans has not been determined. The purpose of this study was to determine the reproducibility of (18)F-FLT microPET studies.C6 rat glioma xenografts were implanted into nude mice (n = 9) and grown to mean diameters of 5-17 mm for approximately 2 wk. A 10-min acquisition was performed on a microPET scanner approximately 1 h after (18)F-FLT (1.9-7.4 MBq [50-200 muCi]) was injected via the tail vein. A second microPET scan was performed approximately 6 h later on the same day after reinjection of (18)F-FLT to assess for reproducibility. Most of the mice were studied twice within the same week (for a total of 17 studies). Images were analyzed by drawing an ellipsoidal region of interest (ROI) around the tumor xenograft activity. Percentage injected dose per gram (%ID/g) values were calculated from the mean activity in the ROIs. Coefficients of variation and differences in %ID/g values between studies from the same day were calculated to determine the reproducibility after subtraction of the estimated residual tumor activity from the first (18)F-FLT injection.The coefficient of variation (mean +/- SD) for %ID/g values between (18)F-FLT microPET scans performed 6 h apart on the same day was 14% +/- 10%. The difference in %ID/g values between scans was -0.06% +/- 1.3%. Serum thymidine levels were mildly correlated with %ID/g values (R(2) = 0.40). Tumor size, mouse body weight, injected dose, and fasting state did not contribute to the variability of the scans; however, consistent scanning parameters were necessary to ensure accurate studies, in particular, controlling body temperature, the time of imaging after injection, and the ROI size.(18)F-FLT microPET mouse tumor xenograft studies are reproducible with moderately low variability. Serial studies may be performed to assess for significant changes in therapy response or for preclinical drug development.

  View details for Web of Science ID 000233095800017

  View details for PubMedID 16269599

• Small animal imaging center design: The facility at the UCLA Crump Institute for Molecular Imaging MOLECULAR IMAGING AND BIOLOGY Stout, D. B., Chatziioannou, A. F., Lawson, T. P., Silverman, R. W., Gambhir, S. S., Phelps, M. E. 2005; 7 (6): 393-402

  Abstract

  The growing number of mouse and rat experiments, coupled with advances in small-animal imaging systems such as microPET, optical, microCAT, microMR, ultrasound and microSPECT, has necessitated a common technical center for imaging small animals.At the UCLA Crump Institute for Molecular Imaging, we have designed and built a facility to support the research interests of a wide range of investigators from multiple disciplines. Requirements to satisfy both research and regulatory oversight have been critically examined. Support is provided for investigator training, study scheduling, data acquisition, archiving, image display, and analysis.The center has been in operation for more than 18 months, supporting more than 13,000 individual imaging procedures.We have created a facility that maximizes our resource utilization while providing optimal investigator support, as well as the means to continually improve the quality and diversity of the science by integrating physical and biological sciences.

  View details for DOI 10.1007/s11307-005-0015-2

  View details for Web of Science ID 000234151200003

  View details for PubMedID 16261425

  View details for PubMedCentralID PMC3005624

• Imaging androgen receptor function during flutamide treatment in the LAPC9 xenograft model MOLECULAR CANCER THERAPEUTICS Ilagan, R., Zhang, L. Q., Pottratz, J., Le, K., Salas, S., Iyer, M., Wu, L., Gambhir, S. S., Carey, M. 2005; 4 (11): 1662-1669

  Abstract

  The current understanding of the response of androgen receptor to pharmacologic inhibitors in prostate cancer is derived primarily from serum prostate-specific antigen (PSA) levels. In this study, we test whether a novel androgen receptor-specific molecular imaging system is able to detect the action of the antiandrogen flutamide on androgen receptor function in xenograft models of prostate cancer. Adenoviruses bearing an optical imaging cassette containing an androgen receptor-responsive two-step transcriptional amplification system were injected into androgen-dependent and hormone-refractory tumors of animals undergoing systemic time-controlled release of the antiandrogen flutamide. Imaging of tumors with a cooled charge-coupled device camera revealed that the response of AdTSTA to flutamide is more sensitive and robust than serum PSA measurements. Flutamide inhibits the androgen signaling pathway in androgen-dependent but not refractory tumors. Analysis of androgen receptor and RNA polymerase II binding to the endogenous PSA gene by chromatin immunoprecipitation revealed that flutamide treatment and androgen withdrawal have different molecular mechanisms. The application of imaging technology to study animal models of cancer provides mechanistic insight into antiandrogen targeting of androgen receptor during disease progression.

  View details for DOI 10.1158/1535-7163.MCT-05-0197

  View details for Web of Science ID 000233265900003

  View details for PubMedID 16275987

• Near-infrared fluorescent RGD peptides for optical imaging of integrin alpha(v)beta 3 expression in living mice BIOCONJUGATE CHEMISTRY Cheng, Z., Wu, Y., Xiong, Z. M., Gambhir, S. S., Chen, X. Y. 2005; 16 (6): 1433-1441

  Abstract

  Near-infrared fluorescence optical imaging is a powerful technique for studying diseases at the molecular level in preclinical models. We recently reported that monomeric RGD peptide c(RGDyK) conjugated to the NIR fluorescent dye specifically targets integrin receptor both in cell culture and in living subjects. In this report, Cy5.5-conjugated mono-, di-, and tetrameric RGD peptides were evaluated in a subcutaneous U87MG glioblastoma xenograft model in order to investigate the effect of multimerization of RGD peptide on integrin avidity and tumor targeting efficacy. The binding affinities of Cy5.5-conjugated RGD monomer, dimer, and tetramer for alpha(v)beta(3) integrin expressed on U87MG cell surface were determined to be 42.9 +/- 1.2, 27.5 +/- 1.2, and 12.1 +/- 1.3 nmol/L, respectively. All three peptide-dye conjugates had integrin specific uptake both in vitro and in vivo. The subcutaneous U87MG tumor can be clearly visualized with each of these three fluorescent probes. Among them, tetramer displayed highest tumor uptake and tumor-to-normal tissue ratio from 0.5 to 4 h postinjection. Tumor-to-normal tissue ratio for Cy5.5-conjugated RGD monomer, dimer, and tetramer were found to be 3.18 +/- 0.16, 2.98 +/- 0.05, and 3.63 +/- 0.09, respectively, at 4 h postinjection. These results suggest that Cy5.5-conjugated monomeric, dimeric, and tetrameric RGD peptides are all suitable for integrin expression imaging. The multmerization of RGD peptide results in moderate improvement of imaging characteristics of the tetramer, compared to that of the monomer and dimeric counterparts.

  View details for DOI 10.1021/bc0501698

  View details for Web of Science ID 000233393800015

  View details for PubMedID 16287239

• Tracking embryonic stem cell transplant in the heart using a novel triple fusion reporter gene 78th Annual Scientific Session of the American-Heart-Association Cao, F., Lin, S., Krishnan, M., Drukker, M. E., Patel, M. R., Ray, P., Zhang, X. Z., Chen, X. Y., Gambhir, S. S., Weissman, I., Wu, J. C. LIPPINCOTT WILLIAMS & WILKINS. 2005: U144–U145

  View details for Web of Science ID 000232956400464

• Adenoviral human BCL-2 attenuates early cell death following cardiomyoblast transplantation into ischemic rat hearts Kutschka, Chen, I. Y., Kofidis, T., Zwierzchoniewska, M., Arai, T., von Degenfeld, G., Sheikh, A. Y., Hendry, S. L., Lebl, D. R., Cooke, D. T., Gambhir, S. S., Robbins, R. C. LIPPINCOTT WILLIAMS & WILKINS. 2005: U504

  View details for Web of Science ID 000232956403068

• In-vivo non-invasive imaging of mesenchymal stem cells after adenoviral or retroviral infection with the herpes simplex virus type-1 thymidine kinase reporter gene Roelants, Bertrand, L., Havaux, Labar, D., Bausart, R., Bol, A., Tabilio, A., Gambhir, S. S., Di Ianni, M., Vanoverschelde, J. L. LIPPINCOTT WILLIAMS & WILKINS. 2005: U824–U825

  View details for Web of Science ID 000232956405210

• Collagen matrices enhance survival of embryonic cardiomyoblasts following transplantation into ischemic rat hearts Kutschka, Kofidis, T., Chen, I. Y., Arai, T., Sheikh, A. Y., Hendry, S. L., Pearl, J., Hoyt, G., Connolly, A., Yang, P. C., Gambhir, S. S., Robbins, R. C. LIPPINCOTT WILLIAMS & WILKINS. 2005: U805

  View details for Web of Science ID 000232956405127

• microPET imaging of glioma integrin alpha(V)beta(3) expression using Cu-64-labeled tetrameric RGD peptide JOURNAL OF NUCLEAR MEDICINE Wu, Y., Zhang, X. Z., Xiong, Z. M., Cheng, Z., Fisher, D. R., Liu, S., Gambhir, S. S., Chen, X. Y. 2005; 46 (10): 1707-1718

  Abstract

  Integrin alpha(v)beta(3) plays a critical role in tumor-induced angiogenesis and metastasis and has become a promising diagnostic indicator and therapeutic target for various solid tumors. Radiolabeled RGD peptides that are integrin specific can be used for noninvasive imaging of integrin expression level as well as for integrin-targeted radionuclide therapy.In this study we developed a tetrameric RGD peptide tracer (64)Cu-DOTA-E{E[c(RGDfK)](2)}(2) (DOTA is 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid) for PET imaging of integrin alpha(v)beta(3) expression in female athymic nude mice bearing the subcutaneous UG87MG glioma xenografts.The RGD tetramer showed significantly higher integrin binding affinity than the corresponding monomeric and dimeric RGD analogs, most likely due to a polyvalency effect. The radiolabeled peptide showed rapid blood clearance (0.61 +/- 0.01 %ID/g at 30 min and 0.21 +/- 0.01 %ID/g at 4 h after injection, respectively [%ID/g is percentage injected dose per gram]) and predominantly renal excretion. Tumor uptake was rapid and high, and the tumor washout was slow (9.93 +/- 1.05 %ID/g at 30 min after injection and 4.56 +/- 0.51 %ID/g at 24 h after injection). The metabolic stability of (64)Cu-DOTA-E{E[c(RGDfK)](2)}(2) was determined in mouse blood, urine, and liver and kidney homogenates at different times after tracer injection. The average fractions of intact tracer in these organs at 1 h were approximately 70%, 58%, 51%, and 26%, respectively. Noninvasive microPET studies showed significant tumor uptake and good contrast in the subcutaneous tumor-bearing mice, which agreed well with the biodistribution results. Integrin alpha(v)beta(3) specificity was demonstrated by successful blocking of tumor uptake of (64)Cu-DOTA-E{E[c(RGDfK)](2)}(2) in the presence of excess c(RGDyK) at 1 h after injection. The highest absorbed radiation doses determined for the human reference adult were received by the urinary bladder wall (0.262 mGy/MBq), kidneys (0.0296 mGy/MBq), and liver (0.0242 mGy/MBq). The average effective dose resulting from a single (64)Cu-DOTA-E{E[c(RGDfK)](2)}(2) injection was estimated to be 0.0164 mSv/MBq.The high integrin and avidity and favorable biokinetics make (64)Cu-DOTA-E{E[c(RGDfK)](2)}(2) a promising agent for peptide receptor radionuclide imaging and therapy of integrin-positive tumors.

  View details for Web of Science ID 000232452700033

  View details for PubMedID 16204722

• Noninvasive imaging of protein-protein interactions from live cells and living subjects using bioluminescence resonance energy transfer FASEB JOURNAL De, A., Gambhir, S. S. 2005; 19 (12): 2017-?

  Abstract

  This study demonstrates a significant advancement of imaging of a distance-dependent physical process, known as the bioluminescent resonance energy transfer (BRET2) signal in living subjects, by using a cooled charge-coupled device (CCD) camera. A CCD camera-based spectral imaging strategy enables simultaneous visualization and quantitation of BRET signal from live cells and cells implanted in living mice. We used the BRET2 system, which utilizes Renilla luciferase (hRluc) protein and its substrate DeepBlueC (DBC) as an energy donor and a mutant green fluorescent protein (GFP2) as the acceptor. To accomplish this objective in this proof-of-principle study, the donor and acceptor proteins were fused to FKBP12 and FRB, respectively, which are known to interact only in the presence of the small molecule mediator rapamycin. Mammalian cells expressing these fusion constructs were imaged using a cooled-CCD camera either directly from culture dishes or by implanting them into mice. By comparing the emission photon yields in the presence and absence of rapamycin, the specific BRET signal was determined. The CCD imaging approach of BRET signal is particularly appealing due to its capacity to seamlessly bridge the gap between in vitro and in vivo studies. This work validates BRET as a powerful tool for interrogating and observing protein-protein interactions directly at limited depths in living mice.

  View details for DOI 10.1096/fj.05-4628fje

  View details for Web of Science ID 000232991100029

• Effects of epigenetic modulation on reporter gene expression: implications for stem cell imaging FASEB JOURNAL Krishnan, M., Park, J. M., Cao, F., Wang, D. X., Paulmurugan, R., Tseng, J. R., Gonzalgo, M. L., Gambhir, S. S., Wu, J. C. 2005; 19 (12): 106-?

  Abstract

  Tracking stem cell localization, survival, differentiation, and proliferation after transplantation in living subjects is essential for understanding stem cell biology and physiology. In this study, we investigated the long-term stability of reporter gene expression in an embryonic rat cardiomyoblast cell line and the role of epigenetic modulation on reversing reporter gene silencing. Cells were stably transfected with plasmids carrying cytomegalovirus promoter driving firefly luciferase reporter gene (CMV-Fluc) and passaged repeatedly for 3-8 months. Within the highest expressor clone, the firefly luciferase activity decreased progressively from passage 1 (843+/-28) to passage 20 (250+/-10) to passage 40 (44+/-3) to passage 60 (3+/-1 RLU/microg; P<0.05 vs. passage 1). Firefly luciferase activity was maximally rescued by treatment with 5-azacytidine (DNA methyltransferase inhibitor) compared with trichostatin A (histone deacetylase inhibitor) and retinoic acid (transcriptional activator; P<0.05). Increasing dosages of 5-azacytidine treatment led to higher levels of firefly luciferase mRNA (RT-PCR) and protein (Western blots) and inversely lower levels of methylation in the CMV promoter (DNA nucleotide sequence). These in vitro results were extended to in vivo bioluminescence imaging (BLI) of cell transplant in living animals. Cells treated with 5-azacytidine were monitored for 2 wk compared with 1 wk for untreated cells (P<0.05). These findings should have important implications for reporter gene-based imaging of stem cell transplantation.

  View details for DOI 10.1096/fj.05-4551fje

  View details for Web of Science ID 000232991100011

  View details for PubMedCentralID PMC3625424

• Micro-PET/CT monitoring of herpes thymidine kinase suicide gene therapy in a prostate cancer xenograft: the advantage of a cell-specific transcriptional targeting approach. Molecular imaging Johnson, M., Sato, M., Burton, J., Gambhir, S. S., Carey, M., Wu, L. 2005; 4 (4): 463-472

  Abstract

  Cancer gene therapy based on tissue-restricted expression of cytotoxic gene should achieve superior therapeutic index over an unrestricted method. This study compared the therapeutic effects of a highly augmented, prostate-specific gene expression method to a strong constitutive promoter-driven approach. Molecular imaging was coupled to gene therapy to ascertain real-time therapeutic activity. The imaging reporter gene (luciferase) and the cytotoxic gene (herpes simplex thymidine kinase) were delivered by adenoviral vectors injected directly into human prostate tumors grafted in SCID mice. Serial bioluminescence imaging, positron emission tomography, and computed tomography revealed restriction of gene expression to the tumors when prostate-specific vector was employed. In contrast, administration of constitutive active vector resulted in strong signals in the liver. Liver serology, tissue histology, and frail condition of animals confirmed liver toxicity suffered by the constitutive active cohorts, whereas the prostate-targeted group was unaffected. The extent of tumor killing was analyzed by apoptotic staining and human prostate marker (prostate-specific antigen). Overall, the augmented prostate-specific expression system was superior to the constitutive approach in safeguarding against systemic toxicity, while achieving effective tumor killing. Integrating noninvasive imaging into cytotoxic gene therapy will provide a useful strategy to monitor gene expression and therapeutic efficacy in future clinical protocols.

  View details for PubMedID 16285908

• Pet Imaging of HSV1-tk Transgene Expression in Cancer Patients Boan, J. F., Penuelas, I., Sangro, B., Mazzolini, G., Ruiz, M., Marti-Climent, J. M., Barrio, J. R., Prieto, J., Gambhir, S. S., Richter, J. A. SPRINGER. 2005: S79

  View details for Web of Science ID 000208745100285

• Novel fusion protein approach for efficient high-throughput screening of small molecule-mediating protein-protein interactions in cells and living animals CANCER RESEARCH Paulmurugan, R., Gambhir, S. S. 2005; 65 (16): 7413-7420

  Abstract

  Networks of protein interactions execute many different intracellular pathways. Small molecules either synthesized within the cell or obtained from the external environment mediate many of these protein-protein interactions. The study of these small molecule-mediated protein-protein interactions is important in understanding abnormal signal transduction pathways in a variety of disorders, as well as in optimizing the process of drug development and validation. In this study, we evaluated the rapamycin-mediated interaction of the human proteins FK506-binding protein (FKBP12) rapamycin-binding domain (FRB) and FKBP12 by constructing a fusion of these proteins with a split-Renilla luciferase or a split enhanced green fluorescent protein (split-EGFP) such that complementation of the reporter fragments occurs in the presence of rapamycin. Different linker peptides in the fusion protein were evaluated for the efficient maintenance of complemented reporter activity. This system was studied in both cell culture and xenografts in living animals. We found that peptide linkers with two or four EAAAR repeat showed higher protein-protein interaction-mediated signal with lower background signal compared with having no linker or linkers with amino acid sequences GGGGSGGGGS, ACGSLSCGSF, and ACGSLSCGSFACGSLSCGSF. A 9 +/- 2-fold increase in signal intensity both in cell culture and in living mice was seen compared with a system that expresses both reporter fragments and the interacting proteins separately. In this fusion system, rapamycin induced heterodimerization of the FRB and FKBP12 moieties occurred rapidly even at very lower concentrations (0.00001 nmol/L) of rapamycin. For a similar fusion system employing split-EGFP, flow cytometry analysis showed significant level of rapamycin-induced complementation.

  View details for DOI 10.1158/0008-5472.CAN-05-0588

  View details for Web of Science ID 000231188600049

  View details for PubMedID 16103094

• Optimizing radiolabeled engineered anti-p185(HER2) antibody fragments for in vivo imaging CANCER RESEARCH Olafsen, T., Kenanova, V. E., Sundaresan, G., Anderson, A. L., Crow, D., Yazaki, P. J., Li, L., Press, M. F., Williams, L. E., Wong, J. Y., Raubitschek, A. A., Shively, J. E., Wu, A. M. 2005; 65 (13): 5907-5916

  Abstract

  We have recently described the in vivo properties of an iodinated anti-p185HER2 engineered antibody fragment [minibody (scFv-C(H)3)2; 80 kDa], made from the internalizing 10H8 monoclonal antibody. Although the 10H8 minibody showed excellent binding to the target in vitro, only modest tumor uptake [5.6 +/- 1.7% injected dose per gram (ID/g) of tissue] was achieved in nude mice bearing MCF7/HER2 breast cancer tumors. Here, in an attempt to improve targeting, the 10H8 minibody was conjugated to 1,4,7,10-tetraazacyclododecane-N, N', N'', N'''-tetraacetic acid (DOTA), radiometal labeled, and evaluated in vivo. The tumor uptake of 111In-DOTA 10H8 minibody was 5.7 +/- 0.1% ID/g, similar to the radioiodinated 10H8 minibody. However, in addition to the expected liver clearance, the kidneys had unexpectedly high activity (34.0 +/- 4.0% ID/g). A minibody derived from a second anti-p185(HER2) antibody (trastuzumab; hu4D5v8) was also made. Tumor uptakes, evaluated by quantitative microPET using 64Cu-DOTA hu4D5v8 minibody, were 4.2 +/- 0.5% ID/g. Furthermore, in non-tumor-bearing mice, 111In-DOTA hu4D5v8 minibody exhibited similar elevated uptake in the kidneys (28.4 +/- 6.5% ID/g). Immunohistochemical staining of kidneys from non-tumor-bearing mice showed strong specific staining of the proximal tubules, and Western blot analysis of kidney lysate confirmed the presence of cross-reactive antigen. To further improve tumor uptake and normal tissue distribution, a larger hu4D5v8 fragment [(scFv-C(H)2-C(H)3)2; 105 kDa] was made, engineered to exhibit rapid clearance kinetics. This fragment, when evaluated by microPET, exhibited improved tumor targeting (12.2 +/- 2.4% ID/g) and reduced kidney uptake (13.1 +/- 1.5% ID/g). Thus, by manipulating the size and format of anti-p185(HER2) antibody fragments, the kidney activity was reduced and high or low expression of p185HER2 in xenografts could be distinguished by microPET imaging.

  View details for Web of Science ID 000230165000055

  View details for PubMedID 15994969

• Noninvasive imaging of ex vivo intracoronarily delivered nonviral therapeutic transgene expression in heart MOLECULAR THERAPY Sen, L., Gambhir, S. S., Furukawa, H., Stout, D. B., Lam, A. L., Laks, H., Cui, G. 2005; 12 (1): 49-57

  Abstract

  We developed a clinically applicable approach for noninvasive monitoring of reporter-therapeutic linked gene expression in the whole heart of large animals using PET imaging and further validated the efficacy and cardiac adverse effects of reporter-therapeutic linked gene transfer in a rabbit cervical heterotopic functional heart transplant model. Cationic liposome complexed with a vector containing a herpes simplex virus type 1 mutant thymidine kinase (HSV1-sr39tk) as the reporter gene and a recombinant human immunosuppressive cytokine, interleukin-10 (hIL-10), as the therapeutic gene was ex vivo intracoronarily delivered into cardiac allografts before implantation. Long-term HSV1-sr39tk and hIL-10 transgene and protein overexpression associated with myocardial PET reporter probe 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]FHBG) accumulation was observed in the allografts. The expression of the HSV1-sr39tk gene was significantly correlated with the hIL-10 gene expression and the total myocardial [18F]FHBG accumulation quantified as a percentage of intravenously injected [18F]FHBG dose. A homogeneous distribution of [18F]FHBG accumulation was seen in the whole heart similar to the distribution of [18F]fluorodeoxyglucose, a PET glucose metabolism probe. The immunosuppressive therapeutic efficacy remained the same in allografts treated with reporter-therapeutic linked gene and therapeutic gene only. No cardiac adverse effect was found. Our results demonstrate for the first time that PET reporter-therapeutic linked gene imaging is applicable for noninvasively monitoring ex vivo intracoronarily delivered therapeutic transgene expression in the whole heart.

  View details for DOI 10.1016/j.ymthe.2005.03.004

  View details for Web of Science ID 000230282200012

  View details for PubMedID 15963920

• Comparison of [C-14]FMAU, [H-3]FEAU, [C-14]FIAU, and [H-3]PCV for monitoring reporter gene expression of wild type,and mutant herpes simplex virus type 1 thymidine kinase in cell culture MOLECULAR IMAGING AND BIOLOGY Kang, K. W., Min, J. J., Chen, X. Y., Gambhir, S. S. 2005; 7 (4): 296-303

  Abstract

  To assess the optimal reporter probe/reporter gene combination for monitoring herpes simplex virus type 1 thymidine kinase (HSV1-tk) gene expression, we compared the cellular uptake of 1-(2'-fluoro-2'-deoxy-D-arabinofuranosyl)-5-methyluracil (FMAU), 2'-fluoro-2'-deoxyarabinofuranosyl-5-ethyluracil (FEAU), 2'-fluoro-2'-deoxy-beta-D-arabinofuranosyl-5-iodouracil (FIAU) and penciclovir (PCV) in both HSV1-tk and HSV1-sr39tk expressing cells.For stably transfected cell studies, C6 rat glioma cells, C6 HSV1-tk transfectant, C6 mutant HSV1-sr39tk transfectant, rat Morris hepatoma cells (MH3924A), and MH3924A HSV1-tk transfectant cells were used. For adenoviral infection studies, C6 rat glioma cells were exposed to serial titers of AdCMV-HSV1-tk, AdCMV-HSV1-sr39tk, or AdCMV-fluc for 24 hours. These cells were incubated with [(14)C]FMAU, [(3)H]FEAU, [(14)C]FIAU, and [(3)H]PCV, and cellular uptake of radioactivity was measured.[(3)H]FEAU exhibited the highest or second highest accumulation and the most selectivity regardless of the mode of gene transfer for both HSV1-tk and mutant HSV1-sr39tk reporter genes.This combination of high accumulation and high selectivity for both HSV1-tk and HSV1-sr39tk makes suitably radiolabeled FEAU a promising candidate as a radiotracer for imaging HSV1-tk/HSV1-sr39tk gene expression in living subjects.

  View details for DOI 10.1007/s11307-005-0010-7

  View details for Web of Science ID 000233422000005

  View details for PubMedID 16041591

• Bioluminescent imaging of melanoma in live mice JOURNAL OF INVESTIGATIVE DERMATOLOGY Craft, N., Bruhn, K. W., Nguyen, B. D., PRINS, R., Liau, L. M., Collisson, E. A., De, A., Kolodney, M. S., Gambhir, S. S., Miller, J. F. 2005; 125 (1): 159-165

  Abstract

  Melanoma is highly resistant to conventional chemotherapeutic agents and novel therapeutic approaches are needed. Current animal models of melanoma in animals are sub-optimal. The most commonly used homograft model is the B16 mouse melanoma. Evaluation of potential melanoma therapies with this model is limited by the inaccuracy of caliper measurement of subcutaneous tumors, of counting lung nodules in metastasis models, and the indirect nature of "survival" curves when studying brain metastases. We have developed and characterized an accurate, sensitive, and reproducible bioluminescent B16 melanoma model that allows for serial, real-time analyses of tumor burden in live mice. We demonstrate that this model is applicable to subcutaneous tumors, lung metastases, and intracranial tumors and offers a solution to many of the limitations of previous models. As proof of principle, we use this model to show the efficacy of a live, Listeria monocytogenes vaccine expressing the melanoma antigen tyrosinase-related protein-2 to protect mice against intravenous B16 melanoma challenge. Additionally, we extend our approach to include the human A375 melanoma model and are able to show in vivo differences between sub-lines with varying metastatic potential. These models represent an accurate and reproducible means for in vivo melanoma monitoring in preclinical studies.

  View details for DOI 10.1111/j.0022-202X.2005.23759.x

  View details for Web of Science ID 000230342000026

  View details for PubMedID 15982316

• Positron emission tomography imaging of adenoviral-mediated transgene expression in liver cancer patients GASTROENTEROLOGY Penuelas, I., Mazzolini, G., Boan, J. F., SANGRO, B., Marti-Climent, J. P., Ruiz, M., Ruiz, J., Satyamurthy, N., Qian, C., Barrio, J. R., Phelps, M. E., Richter, J. A., Gambhir, S. S., Prieto, J. 2005; 128 (7): 1787-1795

  Abstract

  In gene-therapy protocols, imaging of gene expression is needed to evaluate the transduction efficiency of the vector, its tissue distribution, and the duration of transgene expression and to assess the feasibility of repeated vector administration.We have used positron emission tomography with a fluorine-18-labeled penciclovir analogue to monitor thymidine kinase gene expression after intratumoral injection of a first-generation recombinant adenovirus in patients with hepatocellular carcinoma. Patients were enrolled in a pilot clinical trial and treated with escalating doses of the vector. Two days after adenovirus inoculation, transgene expression was evaluated during the first hours after administration of the radiotracer both on the treated lesion and on a whole-body basis.Transgene expression in the tumor was dependent on the injected dose of the adenovirus and was detectable in all patients who received > or = 10(12) viral particles. However, when the study was repeated 9 days after vector injection, no expression could be observed. It is interesting to note that no specific expression of the transgene could be detected in distant organs or in the surrounding cirrhotic tissue in any of the cases studied.Our findings show the real possibility of imaging transgene expression in humans by using viral vectors. We show that hepatocarcinoma is a permissive tumor for adenoviral infection and that the nontumoral cirrhotic liver is spared from transduction when the vector is administered by intratumoral injection. These results show that positron emission tomography imaging may help in the design of gene-therapy strategies and in the clinical assessment of new-generation vectors.

  View details for DOI 10.1053/j.gastro.2005.03.024

  View details for Web of Science ID 000229662900005

  View details for PubMedID 15940613

• Comparison between PET and bioluminescence imaging for quantitative assessment of tumor burden 47th Annual Meeting of the American-Association-of-Physicists-in-Medicine Maxim, P., Thorndyke, B., Boyer, A., Contag, C., Gambhir, S., Xing, L. AMER ASSOC PHYSICISTS MEDICINE AMER INST PHYSICS. 2005: 2134–34

  View details for Web of Science ID 000229908601456

• Functionality of androgen receptor-based gene expression imaging in hormone refractory prostate cancer CLINICAL CANCER RESEARCH Sato, M., Johnson, M., Zhang, L. Q., Gambhir, S. S., Carey, M., Wu, L. 2005; 11 (10): 3743-3749

  Abstract

  A highly augmented, prostate-specific two-step transcriptional amplification (TSTA) method was developed with the ultimate goal of delivering an effective and safe gene-based treatment to prostate cancer patients. Because very limited treatment options are available for recurrent hormone refractory prostate cancer (HRPC), it is imperative to assess whether the prostate-specific antigen (PSA) promoter-based TSTA gene therapy will be functional in HRPC.We tested the TSTA-driven adenovirus vector on three androgen-dependent and six HRPC models. Real-time gene expression was monitored by both optical imaging and the combined modality of positron emission tomography (PET) and computed tomography.The TSTA-driven firefly luciferase expressing adenoviral vector was active in all androgen receptor (AR)-expressing HRPC models, but inactive in AR- and PSA-negative lines. Interestingly, the TSTA-mediated gene expression was induced by hydrocortisone in MDA PCa 2b, a cell line with mutated AR that possesses altered ligand specificity. In animal models, the TSTA-mediated optical signal was more robust in the HRPC than androgen-dependent tumors. In a parallel trend, a TSTA vector that expresses the herpes simplex virus thymidine kinase PET reporter gene also displayed more robust PET signal in the HRPC tumor.The activity of TSTA system is AR dependent and it recapitulates the functional status of endogenous AR. These data support the conclusion that AR function is activated in HRPC despite castrated levels of androgen. Together with the fact that majority of recurrent prostate cancers express AR and PSA, we foresee that the TSTA approach can be a promising gene therapy strategy for the advanced stages of prostate cancer.

  View details for Web of Science ID 000229086600019

  View details for PubMedID 15897571

  View details for PubMedCentralID PMC2821218

• A new strategy to screen molecular imaging probe uptake in cell culture without radiolabeling using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry JOURNAL OF NUCLEAR MEDICINE Cheng, Z., Winant, R. C., Gambhir, S. S. 2005; 46 (5): 878-886

  Abstract

  Numerous new molecular targets for diseases are rapidly being identified and validated in the postgenomic era, urging scientists to explore novel techniques for accelerating molecular probe development. In this study, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was investigated as a potential tool for high-throughput screening and characterization of molecular imaging probes. Specifically, MALDI-TOF-MS was used to screen a small library of phosphonium cations for their ability to accumulate in cells.C6 cells incubated with phosphonium cations at room temperature were collected and lysed for experiments. Calibration curves for the internal standard, methyltriphenyl phosphonium, and for tetraphenylphosphonium bromide (TPP) and other phosphonium cations were first established. The time course of TPP uptake by C6 cells was then quantified using both MALDI-TOF-MS and liquid scintillation counting with (3)H-TPP. In addition, MALDI-TOF-MS was used to screen a library of 8 phosphonium cations and subsequently rank their ability to penetrate membranes and accumulate in cells. Finally, the accumulation of 4-fluorophenyltriphenyl phosphonium (FTPP) in the membrane potential-modulated cells was also measured by MALDI-TOF-MS.MALDI-TOF-MS spectra clearly revealed that TPP was easily identified from cell lysates even as early as 10 min after incubation and that levels as low as 0.11 fmol of TPP per cell could be detected, suggesting the high sensitivity of this technique. The time course of TPP influx determined by both MALDI-TOF-MS and radioactivity counting showed no statistically significant difference (P > 0.05 for all time points). These data validated MALDI-TOF-MS as an alternative approach for accurately measuring uptake of phosphonium cations by cells. TPP and FTPP demonstrated greater accumulation in cells than did the other cations evaluated in this study. Furthermore, uptake profiles suggested that FTPP preserves the membrane potential-dependent uptake property of TPP in cell cultures. Taken together, these data justify further synthesis and evaluation of (18)F-FTPP as a molecular probe for imaging mitochondrial dysfunction.These results demonstrate that MALDI-TOF-MS is a powerful analytic tool for rapid screening and characterization of phosphonium cations as molecular probes. This technique can potentially be applied to the evaluation of other imaging probes or drugs and thus may facilitate their rational design and development.

  View details for Web of Science ID 000228952400030

  View details for PubMedID 15872363

• Imaging protein-protein interactions in living subjects TRAC-TRENDS IN ANALYTICAL CHEMISTRY Paulmurugan, R., Ray, P., De, A., CHAN, C. T., Gambhir, S. S. 2005; 24 (5): 446-458

  View details for Web of Science ID 000234203000028

• Noninvasive monitoring of target gene expression by imaging reporter gene expression in living animals using improved bicistronic vectors JOURNAL OF NUCLEAR MEDICINE Wang, Y. L., Iyer, M., Annala, A. J., Chappell, S., Mauro, V., Gambhir, S. S. 2005; 46 (4): 667-674

  Abstract

  Indirect, noninvasive imaging of therapeutic gene expression based on levels of reporter gene expression is a powerful tool to devise improved therapeutic strategies in cancer gene therapy. The use of bicistronic vectors carrying internal ribosome entry sites (IRESs) allows the coexpression of multiple gene products from the same promoter but leads to considerable attenuation of the downstream gene. In this study, we describe the use of 10 linked copies of the Gtx (homeodomain protein) IRES (abbreviated as SIRES) in place of the encephalomyocarditis (EMCV) IRES in mediating downstream reporter gene expression in cell culture and in vivo.We constructed several plasmid vectors carrying different upstream and downstream reporter genes (herpes simplex virus type I thymidine kinase [tk], firefly luciferase [fl], and Renilla luciferase [rl]) placed between EMCV IRES and SIRES segments. RL, FL, and TK enzyme activities in N2a, C6, and 293 cells transiently transfected with these vectors were found to be significantly higher for the SIRES vectors than for the EMCV IRES vectors. For in vivo experiments, 4 stably transfected N2a cell lines were implanted in nude mice. The mice were imaged for rl and fl gene expression using a charged-coupled device (CCD) camera. For bioluminescence and microPET imaging of downstream gene expression of fl and tk genes, respectively, mice carrying 4 stably transfected xenografts were imaged using the CCD camera and microPET.In cell culture, using rl as the upstream gene, we demonstrate that the expression of the downstream tk gene is 12-fold greater using SIRES when compared with EMCV IRES. Furthermore, the expression of the 2 genes was highly correlated in N2a cells. In vivo bioluminescence imaging using 4 stably transfected N2a cell lines revealed increasing levels of rl and fl gene expression. Bioluminescence and microPET, respectively, of fl and tk reporter gene expression in nude mice bearing N2a tumor xenografts showed the gene expression mediated by SIRES to be 4- and 8-fold higher, respectively, than EMCV IRES.These findings support the use of SIRES bicistronic vectors for a better assessment of therapeutic gene expression based on reporter gene expression in living subjects.

  View details for Web of Science ID 000228200000022

  View details for PubMedID 15809490

• Pet imaging allows monitoring transgene expression in gene therapy of liver cancer using adenoviral vectors Sangro, B., Penuelas, Mazzolini, G., Boan, J., Marti, J., Ruiz, M., Satyamurthy, N., Barrio, J., Phelps, M., Richter, J., Gambhir, S., Prieto, J. ELSEVIER SCIENCE BV. 2005: 140–41

  View details for Web of Science ID 000229024000382

• FDG-PET and beyond: Molecular breast cancer imaging JOURNAL OF CLINICAL ONCOLOGY Quon, A., Gambhir, S. S. 2005; 23 (8): 1664-1673

  View details for DOI 10.1200/JCO.2005.11.024

  View details for Web of Science ID 000227587200017

  View details for PubMedID 15755974

• Imaging progress of herpes simplex virus type 1 thymidine kinase suicide gene therapy in living subjects with positron emission tomography CANCER GENE THERAPY Yaghoubi, S. S., Barrio, J. R., Namavari, M., Satyamurthy, N., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2005; 12 (3): 329-339

  Abstract

  Molecular imaging of a suicide transgene's expression will aid the development of efficient and precise targeting strategies, and imaging for cancer cell viability may assess therapeutic efficacy. We used the PET reporter probe, 9-(4-[18F]fluoro-3-(hydroxymethyl)butyl)guanine ([18F]FHBG) to monitor the expression of a mutant Herpes Simplex Virus 1 thymidine kinase (HSV1-sr39tk) in C6 glioma tumors implanted subcutaneously in nude mice that were repetitively being treated with the pro-drug Ganciclovir (GCV). [18F]-Fluorodeoxyglucose ([18F]FDG), a metabolic tracer, was used to assess tumor cell viability and therapeutic efficacy. C6 glioma tumors stably expressing the HSV1-sr39tk gene (C6sr39) accumulated [18F]FHBG prior to GCV treatment. Significant declines in C6sr39 tumor volumes and [18F]FHBG and [18F]FDG accumulation were observed following 2 weeks of GCV treatment. However, 3 weeks after halting GCV treatment, the tumors re-grew and [18F]FDG accumulation increased significantly; in contrast, tumor [18F]FHBG concentrations remained at background levels. Therefore, [18F]FHBG can be used to detect tumors expressing HSV1-sr39tk, susceptible to regression in response to GCV exposure, and the effectiveness of GCV therapy in eradicating HSV1-sr39tk-expressing cells can be monitored by [18F]FHBG scanning. [18F]FHBG and [18F]FDG imaging data indicate that exposure of C6sr39 tumors to GCV causes the elimination of [18F]FHBG-accumulating C6sr39 cells and selects for re-growth of tumors unable to accumulate [18F]FHBG.

  View details for DOI 10.1038/sj.cgt.7700795

  View details for Web of Science ID 000227026700013

  View details for PubMedID 15592447

• Firefly luciferase enzyme fragment complementation for imaging in cells and living animals ANALYTICAL CHEMISTRY Paulmurugan, R., Gambhir, S. S. 2005; 77 (5): 1295-1302

  Abstract

  We identified different fragments of the firefly luciferase gene based on the crystal structure of firefly luciferase. These split reporter genes which encode for protein fragments, unlike the fragments currently used for studying protein-protein interactions, can self-complement and provide luciferase enzyme activity in different cell lines in culture and in living mice. The comparison of the fragment complementation associated recovery of firefly luciferase enzyme activity with intact firefly luciferase was estimated for different fragment combinations and ranged from 0.01 to 4% of the full firefly luciferase activity. Using a cooled optical charge-coupled device camera, the analysis of firefly luciferase fragment complementation in transiently transfected subcutaneous 293T cell implants in living mice showed significant detectable enzyme activity upon injecting d-luciferin, especially from the combinations of fragments identified (Nfluc and Cfluc are the N and C fragments of the firefly luciferase gene, respectively): Nfluc (1-475)/Cfluc (245-550), Nfluc (1-475)/Cfluc (265-550), and Nfluc (1-475)/Cfluc (300-550). The Cfluc (265-550) fragment, upon expression with the nuclear localization signal (NLS) peptide of SV40, shows reduced enzyme activity when the cells are cotransfected with the Nfluc (1-475) fragment expressed without NLS. We also proved in this study that the complementing fragments could be efficiently used for screening macromolecule delivery vehicles by delivering TAT-Cfluc (265-550) to cells stably expressing Nfluc (1-475) and recovering signal. These complementing fragments should be useful for many reporter-based assays including intracellular localization of proteins, studying cellular macromolecule delivery vehicles, studying cell-cell fusions, and also developing intracellular phosphorylation sensors based on fragment complementation.

  View details for DOI 10.1021/ac0484777

  View details for Web of Science ID 000227409800021

  View details for PubMedID 15732910

• Non-invasive imaging of a transgenic mouse model using a prostate-specific two-step transcriptional amplification strategy TRANSGENIC RESEARCH Iyer, M., Salazar, F. B., Lewis, X., Zhang, L., Wu, L., Carey, M., Gambhir, S. S. 2005; 14 (1): 47-55

  Abstract

  Non-invasive assessment of transgenic animals using bioluminescence imaging offers a rapid means of evaluating disease progression in animal models of disease. One of the challenges in the field is to develop models with robust expression to image repetitively live intact animals through solid tissues. The prostate-specific antigen (PSA) promoter is an attractive model for studying gene regulation due to its hormonal response and tissue-specificity permitting us to measure signaling events that occur within the native tissues. The use of the GAL4-VP16 activator offers a powerful means to augment gene expression levels driven by a weak promoter. We have used a two-step transcriptional amplification (TSTA) system to develop a transgenic mouse model to investigate the tissue-specificity and developmental regulation of firefly luciferase (fl) gene expression in living mice using bioluminescence imaging. We employed an enhanced prostate-specific promoter to drive the yeast transcriptional activator, GAL4-VP16 (effector). The reporter construct carries five Gal4 binding sites upstream of the fl gene. We generated a transgenic mouse model using a single vector carrying the effector and reporter constructs. The transgenic mice show prostate-specific expression as early as three weeks of age. The bioluminescence signal in the prostate is significantly higher than in other organs. We also demonstrate that blocking androgen availability can downregulate the fl expression in the prostate. The transgenic mice display normal physical characteristics and developmental behavior, indicating that the high level of GAL4 driven expression is well tolerated. These findings suggest that the GAL4-VP16 transactivator can be used to amplify reporter gene expression from a relatively weak promoter in a transgenic mouse model. The transgenic TSTA model in conjunction with other transgenic cancer models should also help to detect and track malignancies. The strategies developed will be useful for transgenic research in general by allowing for amplified tissue specific gene expression.

  View details for DOI 10.1007/s11248-004-2836-1

  View details for Web of Science ID 000227424200005

  View details for PubMedID 15865048

• Quantum dots for live cells, in vivo imaging, and diagnostics SCIENCE Michalet, X., Pinaud, F. F., Bentolila, L. A., Tsay, J. M., Doose, S., Li, J. J., Sundaresan, G., Wu, A. M., Gambhir, S. S., Weiss, S. 2005; 307 (5709): 538-544

  Abstract

  Research on fluorescent semiconductor nanocrystals (also known as quantum dots or qdots) has evolved over the past two decades from electronic materials science to biological applications. We review current approaches to the synthesis, solubilization, and functionalization of qdots and their applications to cell and animal biology. Recent examples of their experimental use include the observation of diffusion of individual glycine receptors in living neurons and the identification of lymph nodes in live animals by near-infrared emission during surgery. The new generations of qdots have far-reaching potential for the study of intracellular processes at the single-molecule level, high-resolution cellular imaging, long-term in vivo observation of cell trafficking, tumor targeting, and diagnostics.

  View details for DOI 10.1126/science.1104274

  View details for Web of Science ID 000226694000036

  View details for PubMedID 15681376

  View details for PubMedCentralID PMC1201471

• Tailoring the pharmacokinetics and positron emission tomography imaging properties of anti-carcinoembryonic antigen single-chain Fv-Fc antibody fragments CANCER RESEARCH Kenanova, V., Olafsen, T., Crow, D. M., Sundaresan, G., Subbarayan, M., Carter, N. H., Ikle, D. N., Yazaki, P. J., Chatziioannou, A. F., Gambhir, S. S., Williams, L. E., Shively, J. E., Colcher, D., Raubitschek, A. A., Wu, A. M. 2005; 65 (2): 622-631

  Abstract

  Antibody fragments are recognized as promising vehicles for delivery of imaging and therapeutic agents to tumor sites in vivo. The serum persistence of IgG1 and fragments with intact Fc region is controlled by the protective neonatal Fc receptor (FcRn) receptor. To modulate the half-life of engineered antibodies, we have mutated the Fc-FcRn binding site of chimeric anti-carcinoembryonic antigen (CEA) antibodies produced in a single-chain Fv-Fc format. The anti-CEA T84.66 single-chain Fv-Fc format wild-type and five mutants (I253A, H310A, H435Q, H435R, and H310A/H435Q, Kabat numbering system) expressed well in mammalian cell culture. After purification and characterization, effective in vitro antigen binding was shown by competition ELISA. Biodistribution studies in BALB/c mice using (125)I- and (131)I-labeled fragments revealed blood clearance rates from slowest to fastest as follows: wild-type > H435R > H435Q > I253A > H310A > H310A/H435Q. The terminal half-lives of the mutants ranged from 83.4 to 7.96 hours, whereas that of the wild-type was approximately 12 days. Additionally, (124)I-labeled wild-type, H435Q, I253A, H310A, and H310A/H435Q variants were evaluated in LS174T xenografted athymic mice by small animal positron emission tomography imaging, revealing localization to the CEA-positive xenografts. The slow clearing wild-type and H435Q constructs required longer to localize to the tumor and clear from the circulation. The I253A and H310A fragments showed intermediate behavior, whereas the H310A/H435Q variant quickly localized to the tumor site, rapidly cleared from the animal circulation and produced clear images. Thus, attenuating the Fc-FcRn interaction provides a way of controlling the antibody fragment serum half-life without compromising expression and tumor targeting.

  View details for Web of Science ID 000226334500033

  View details for PubMedID 15695407

• Studying molecular and cellular processes in the intact organism. Progress in drug research. Fortschritte der Arzneimittelforschung. Progrès des recherches pharmaceutiques Gheysens, O., Gambhir, S. S. 2005; 62: 117-150

  View details for PubMedID 16329256

• Non-Invasive Imaging Strategies to Visualize Tissue-Specific Gene Expression Using Transcriptional Amplification Approaches Molecular Imaging of the Lungs Iyer, M., Gambhir, S. S. edited by Schuster, D. P., Blackwell, T. Marcel Dekker, Inc.. 2005: 172–206
• Protein-Protein Interaction Imaging In Living Subjects A Molecular Cloning Manual: Protein-Protein Interactions Paulmurugan, R., Ray, P., De, A., Chan, C., Gambhir, S. S. edited by Golemis, E., Adams, P. Cold Spring Harbor Laboratory Press. 2005; 2: 695–713
• Studying Molecular and Cellular Processes in the Intact Organism Progress in Drug Research Gheysens, O., Gambhir, S. S. edited by Rudin, M. Birkhauser-Verlag AG: Basel. 2005: 117–150
• Molecular Imaging of Gene Products Clinical Nuclear Cardiology: State of the Art and Future Directions Wu, J., Gambhir, S. S. edited by Zaret, B., Beller, G. Mosby, Inc.. 2005: 673–690
• Small Animal Imaging Center Design: The Facility at the UCLA Crump Institute for Molecular Imaging Molecular Imaging and Biology Stout, D. B., Chatziioannou, A. F., Lawson, T. P., Silverman, R. W., Gambhir, S. S., Phelps, M. E. 2005: 393-402
• Visualiztion of a Primary Anti-Tumor Immune REsponse by Positron Emission Tomography Proceedings of the National Academy of Sciences (USA) Shu, C., Guo, S., Kim , Y., Shelly, S., Nijagal, A., Ray, P., Gambhir, S. S., Radu, C., Witte, O. 2005: 17412-17417
• MicroPET/CT Monitoring of Herpes Thymidine Kinase Suicide Gene Therapy in a Prostate Cancer Xenograft: The Advantage of a Cell-Specific Transcriptional Targeting Approach Molecular Imaging Johnson, M., Sato, M., Burton, J., Gambhir, S. S., Carey, M., Wu, L. 2005: 463-472
• A New Strategy to Screen Molecular Imaging Probe Uptake in Cell Culture without Radiolabeling using Matrix Assisted Laser Desorption/Ionization time of Flight Mass Spectrometry A New Strategy to Screen Molecular Imaging Probe Uptake in Cell Culture without Radiolabeling using Matrix Assisted Laser Desorption/Ionization time of Flight Mass Spectrometry Cheng, Z. 2005; 46: 878-886
• Multimodality Radionuclide, Fluorescence, and Bioluminescence Small-Animal Imaging. IEEE Park J, Gambhir SS 2005; 93: 771-783
• Quantum Dots for Molecular Imaging and Cancer Medicine. Discovery Medicine Bentolila LA, Michalet X, Pinaud Ff, Tsay JM, Doose S, Li JJ, Sundaresan G, Wu AM, Gambhir SS, Weiss S 2005; 5(26): 213-218
• Comparison between PET and bioluminescence imaging for quantitative assessment of tumor burden 47th Annual Meeting of the American-Society-for-Therapeutic-Radiology-and-Oncology Maxim, P. G., Thorndyke, B. R., Dandekar, M., Mayer, D., Contag, C. H., Gambhir, S. S., Xing, L. ELSEVIER SCIENCE INC. 2005: S490–S490

  View details for Web of Science ID 000232083301348

• Synthesis of (4-[18F]Fluorophenyl) triphenylphosphonium as a Potential Imaging Agent for Mitochondrial Dysfunction. Journal of Labeled Compound and Radiopharmaceuticals Cheng Z, Subbarayan M, Chen X, Gambhir SS 2005; 48: 131-137
• Multimodality Tumor Imaging Targeting Integrin &#945;v&#946;3 BioTechniques Cai W, Gambhir SS, Chen X 2005; 39: S1-S25
• Clinical Molecular Imaging and Therapy-Moving Ahead Together. European Journal of Nuclear Medicine and Molecular Imaging Bengel FM, Gambhir SS. 2005; 32 Suppl 2: S323
• Gene Therapy Imaging in Patients for Oncological Applications. European Journal of Nuclear Medicine and Molecular Imaging Panuelas I, Haberkorn U, Yaghoubi S, Gambhir SS. 2005; 32:Suppl 2: S384-403
• A New Strategy to Screen Molecular Imaging Probe Uptake in Cell Culture without Radiolabeling using Matrix Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry. Journal of Nuclear Medicine Cheng Z, Winant RC, Gambhir SS. 2005; 46: 878-886
• Re: Tetraphenylphosphonium as a novel molecular probe for imaging tumors - Reply JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 2004; 45 (12): 2126

  View details for Web of Science ID 000225927200028

• PET of cardiac transgene expression: Comparison of 2 approaches based on herpesviral thymidine kinase reporter gene JOURNAL OF NUCLEAR MEDICINE Miyagawa, M., Anton, M., Haubner, R., Simoes, M. V., Stadele, C., Erhardt, W., Reder, S., Lehner, T., Wagner, B., Noll, S., Noll, B., Grote, M., Gambhir, S. S., Gansbacher, B., Schwaiger, M., Bengel, F. M. 2004; 45 (11): 1917-1923

  Abstract

  PET of reporter gene expression holds promise for noninvasive monitoring of gene therapy. Previously, 2 approaches based on the herpes simplex virus type 1 thymidine kinase gene (HSV1-tk) have been successfully applied to the heart. Wild-type HSV1-tk was imaged with (124)I-labeled 2'-fluoro-2'-deoxy-5-iodo-1-beta-D-arabinofuranosyl-5-iodouracil (FIAU), and a mutant HSV1-tk (HSV1-sr39tk) was imaged with (18)F-labeled 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine (FHBG). The aim of this study was to compare these 2 combinations with regard to specificity, imaging contrast, and reporter probe kinetics using dynamic PET in small and large animals.Similar titers of adenovirus-expressing wild-type HSV1-tk (Ad(tk)), mutant HSV1-sr39tk (Ad(sr39tk)), or control genes were directly injected into the myocardium of 24 rats and 8 pigs. Two days later, dynamic PET was performed with a clinical scanner during the 120 min after injection of (124)I-FIAU (Ad(tk) animals and controls) or (18)F-FHBG (Ad(sr39tk) animals and controls). Imaging with (13)N-ammonia was performed to identify cardiac regions of interest.In rats, significant cardiac (124)I-FIAU accumulation occurred in images obtained early (10-30 min) after Ad(tk) injection. Because of tracer washout, however, no difference between Ad(tk)-injected animals and controls was seen in the images obtained later. For (18)F-FHBG, specific myocardial accumulation greater than background levels was detected in Ad(sr39tk)-injected animals at early imaging and, in contrast to (124)I-FIAU accumulation, increased over time until the latest imaging (105-120 min). At maximum, cardiac (18)F-FHBG concentration showed a 4.15 +/- 1.65-fold increase compared with controls (105-120 min), and cardiac (124)I-FIAU concentration reached a maximal increase of 1.34 +/- 0.38-fold compared with controls (10-30 min, P = 0.0014). Global cardiac reporter probe kinetics in rats were confirmed by regional myocardial analysis in pig hearts. Transgene expression was specifically visualized by both approaches. The highest target-to-background ratio of (124)I-FIAU in Ad(tk)-infected pig myocardium was 1.50 +/- 0.20, versus 2.64 +/- 0.49 for (18)F-FHBG in Ad(sr39tk)-infected areas (P = 0.01). In vivo results were confirmed by ex vivo counting and autoradiography.Both reporter gene/probe combinations were feasible for noninvasive imaging of cardiac transgene expression in different species. Specific probe kinetics suggest different myocardial handling of pyrimidine (FIAU) and acycloguanosine (FHBG) derivatives. The results favor (18)F-FHBG with mutant HSV1-sr39tk because of continuous accumulation over time and higher imaging contrast.

  View details for Web of Science ID 000225086000026

  View details for PubMedID 15534063

• A tracer kinetic model for F-18-FHBG for quantitating herpes simplex virus type 1 thymidine kinase reporter gene expression in living animals using PET 47th Annual Meeting of the Society-of-Nuclear-Medicine Green, L. A., Nguyen, K., Berenji, B., Iyer, M., Bauer, E., Barrio, J. R., Namavari, M., Satyamurthy, N., Gambhir, S. S. SOC NUCLEAR MEDICINE INC. 2004: 1560–70

  Abstract

  Reporter probe 9-(4-18F-fluoro-3-[hydroxymethyl]butyl)guanine (18F-FHBG) and reporter gene mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) have been used for imaging reporter gene expression with PET. Current methods for quantitating the images using the percentage injected dose per gram of tissue do not distinguish between the effects of probe transport and subsequent phosphorylation. We therefore investigated tracer kinetic models for 18F-FHBG dynamic microPET data and noninvasive methods for determining blood time-activity curves in an adenoviral gene delivery model in mice.18F-FHBG (approximately 7.4 MBq [approximately 200 microCi]) was injected into 4 mice; 18F-FHBG concentrations in plasma and whole blood were measured from mouse heart left ventricle (LV) direct sampling. Replication-incompetent adenovirus (0-2 x 10(9) plaque-forming units) with the E1 region deleted (n = 8) or replaced by HSV1-sr39tk (n = 18) was tail-vein injected into mice. Mice were dynamically scanned using microPET (approximately 7.4 MBq [approximately 200 microCi] 18F-FHBG) over 1 h; regions of interest were drawn on images of the heart and liver. Serial whole blood 18F-FHBG concentrations were measured in 6 of the mice by LV sampling, and 1 least-squares ratio of the heart image to the LV time-activity curve was calculated for all 6 mice. For 2 control mice and 9 mice expressing HSV1-sr39tk, heart image (input function) and liver image time-activity curves (tissue curves) were fit to 2- and 3-compartment models using Levenberg-Marquardt nonlinear regression. The models were compared using an F statistic. HSV1-sr39TK enzyme activity was determined from liver samples and compared with model parameter estimates. For another 3 control mice and 6 HSV1-sr39TK-positive mice, the model-predicted relative percentage of metabolites was compared with high-performance liquid chromatography analysis.The ratio of 18F-FHBG in plasma to whole blood was 0.84 +/- 0.05 (mean +/- SE) by 30 s after injection. The least-squares ratio of the heart image time-activity curve to the LV time-activity curve was 0.83 +/- 0.02, consistent with the recovery coefficient for the partial-volume effect (0.81) based on independent measures of heart geometry. A 3-compartment model best described 18F-FHBG kinetics in mice expressing HSV1-sr39tk in the liver; a 2-compartment model best described the kinetics in control mice. The 3-compartment model parameter, k3, correlated well with the HSV1-sr39TK enzyme activity (r2 = 0.88).18F-FHBG equilibrates rapidly between plasma and whole blood in mice. Heart image time-activity curves corrected for partial-volume effects well approximate LV time-activity curves and can be used as input functions for 2- and 3-compartment models. The model parameter k3 from the 3-compartment model can be used as a noninvasive estimate for HSV1-sr39TK reporter protein activity and can predict the relative percentage of metabolites.

  View details for Web of Science ID 000223809500025

  View details for PubMedID 15347725

• Multimodality imaging of lymphocytic migration using lentiviral-based transduction of a tri-fusion reporter gene MOLECULAR IMAGING AND BIOLOGY Kim, Y. J., Dubey, P., Ray, P., Gambhir, S. S., Witte, O. N. 2004; 6 (5): 331-340

  Abstract

  Previous work showed quantitative imaging of T-cell migration into a tumor site by positron emission tomography (PET), using retroviral transduction of mutated thymidine kinase (sr39TK) reporter genes into immunized T-lymphocytes. PROCEDURES AND RESULTS: In order to improve the sensitivity and flexibility of the imaging analysis, lentivirus, that expressed sr39TK, was used to transduce the lymphocytes that migrated to an immunogenic sarcoma site. In comparison to retrovirally transduced lymphocytes, the lentivirally transduced lymphocytes showed enhanced PET signal when equal numbers of transduced lymphocytes were transferred. Furthermore, in order to utilize multimodality in vivo imaging capability, a tri-fusion reporter gene containing sr39TK, synthetic Renilla luciferase (hRluc), and enhanced green fluorescent protein (eGFP) was inserted into a lentiviral transfer vector. Using the adoptive transfer model, tumor-specific lymphocytic migration was detected by both microPET scan and bioluminescence imaging.The multimodal imaging strategy coupled with lentiviral reporter construct delivery demonstrated here can facilitate future molecular imaging studies.

  View details for DOI 10.1016/j.mibio.2004.06.009

  View details for Web of Science ID 000224370600007

  View details for PubMedID 15380743

• Noninvasive Imaging of enhanced prostate-specific gene expression using a two-step transcriptional amplification-based lentivirus vector MOLECULAR THERAPY Iyer, M., Salazar, F. B., Lewis, X., Zhang, L. Q., Carey, M., Wu, L., Gambhir, S. S. 2004; 10 (3): 545-552

  Abstract

  Noninvasive evaluation of gene transfer to specific cells or tissues will allow for long-term, repetitive monitoring of transgene expression. Tissue-specific promoters that restrict the expression of a transgene to tumor cells play a vital role in cancer gene therapy imaging. In this study, we have developed a third-generation HIV-1-based lentivirus vector carrying a prostate-specific promoter to monitor the long-term, sustained expression of the firefly luciferase (fl) reporter gene in living mice. The fl gene in the transcriptionally targeted vector is driven by an enhanced prostate-specific antigen promoter in a two-step transcriptional amplification (TSTA) system. The efficiency of the lentivirus (LV-TSTA)-mediated gene delivery, cell-type specificity, and persistence of gene expression were evaluated in cell culture and in living mice carrying prostate tumor xenografts. In vivo bioluminescence imaging with a cooled charge-coupled device camera revealed significantly high levels of fl expression in prostate tumors. Injection of LV-TSTA directly into the prostate of male nude mice revealed efficient and long-term fl gene expression in the prostate tissue for up to 3 months. These studies demonstrate the significant potential of TSTA-based lentivirus vectors to confer high levels of tissue-specific gene expression from a weak promoter, while preserving cell-type specificity and the ability to image noninvasively the sustained, long-term expression of reporter genes in living animals.

  View details for DOI 10.1016/j.ymthe.2004.06.118

  View details for Web of Science ID 000224480600019

  View details for PubMedID 15336654

• Molecular imaging of the kinetics of vascular endothelial growth factor gene expression in ischemic myocardium CIRCULATION Wu, J. C., Chen, I. Y., Wang, Y. L., Tseng, J. R., Chhabra, A., Salek, M., Min, J. J., Fishbein, M. C., Crystal, R., Gambhir, S. S. 2004; 110 (6): 685-691

  Abstract

  Angiogenic gene therapy is a promising treatment paradigm for patients with ischemic heart disease. In this study, we used micro-positron emission tomography (microPET) to monitor the transgene expression, function, and effects in a whole-body system.Adenovirus with cytomegalovirus promoter driving an angiogenic gene (vascular endothelial growth factor [VEGF]) linked to a PET reporter gene (herpes simplex virus type 1 mutant thymidine kinase; Ad-CMV-VEGF121-CMV-HSV1-sr39tk) was used to transfect rat embryonic cardiomyoblasts in vitro. Expression of both genes correlated strongly (r=0.98; P<0.001). Afterward, rats underwent ligation of the left anterior descending artery followed by injection of 1x10(10) pfu of Ad-CMV-VEGF121-CMV-HSV1-sr39tk (study; n=35) or Ad-null (control; n=15) at the peri-infarct region. Noninvasive microPET imaging was used to assess the uptake of 9-(4-[18F]-fluoro-hydroxymethylbutyl)guanine ([18F]-FHBG) PET reporter probe by cells expressing the HSV1-sr39tk PET reporter gene. Cardiac transgene expression peaked at day 1 and declined over the next 2 weeks. Repeat adenoviral injections at day 60 yielded no detectable signal. The in vivo reporter gene expression (% injected dose/g of [18F]-FHBG) correlated well with ex vivo gamma counting (r=0.92), myocardial tissue HSV1-sr39TK enzyme activity (r=0.95), and myocardial tissue VEGF level (r=0.94; P<0.001 for all). The VEGF121 isoform induced significant increases in capillaries and small blood vessels. However, the level of neovasculature did not translate into significant improvements in functional parameters such as myocardial contractility by echocardiography, perfusion by nitrogen-13 ammonia imaging, and metabolism by [18F]-fluorodeoxyglucose imaging.Taken together, these findings establish the feasibility of molecular imaging for monitoring angiogenic gene expression with a PET reporter gene and probe noninvasively, quantitatively, and repetitively. The principles demonstrated here can be used to evaluate other therapeutic genes of interest in animal models before future clinical trials are initiated.

  View details for DOI 10.1161/01.CIR.0000138153.02213.22

  View details for Web of Science ID 000223194700008

  View details for PubMedID 15302807

• Molecular imaging of cardiovascular gene products JOURNAL OF NUCLEAR CARDIOLOGY Wu, J. C., Tseng, J. R., Gambhir, S. S. 2004; 11 (4): 491-505

  View details for DOI 10.1016/j.nuclcard.2004.04.004

  View details for Web of Science ID 000223210200015

  View details for PubMedID 15295418

• Novel bidirectional vector strategy for amplification of therapeutic and reporter gene expression HUMAN GENE THERAPY Ray, S., Paulmurugan, R., Hildebrandt, I., Iyer, M., Wu, L., Carey, M., Gambhir, S. S. 2004; 15 (7): 681-690

  Abstract

  Molecular imaging methods have previously been employed to image tissue-specific reporter gene expression by a two-step transcriptional amplification (TSTA) strategy. We have now developed a new bidirectional vector system, based on the TSTA strategy, that can simultaneously amplify expression for both a target gene and a reporter gene, using a relatively weak promoter. We used the synthetic Renilla luciferase (hrl) and firefly luciferase (fl) reporter genes to validate the system in cell cultures and in living mice. When mammalian cells were transiently cotransfected with the GAL4-responsive bidirectional reporter vector and various doses of the activator plasmid encoding the GAL4-VP16 fusion protein, pSV40-GAL4-VP16, a high correlation (r(2) = 0.95) was observed between the expression levels of both reporter genes. Good correlations (r(2) = 0.82 and 0.66, respectively) were also observed in vivo when the transiently transfected cells were implanted subcutaneously in mice or when the two plasmids were delivered by hydrodynamic injection and imaged. This work establishes a novel bidirectional vector approach utilizing the TSTA strategy for both target and reporter gene amplification. This validated approach should prove useful for the development of novel gene therapy vectors, as well as for transgenic models, allowing noninvasive imaging for indirect monitoring and amplification of target gene expression.

  View details for Web of Science ID 000222786900006

  View details for PubMedID 15242528

  View details for PubMedCentralID PMC4153396

• Molecular imaging of gene expression in living subjects by spliceosome-mediated RNA trans-splicing PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Bhaumik, S., Walls, Z., Puttaraju, M., Mitchell, L. G., Gambhir, S. S. 2004; 101 (23): 8693-8698

  Abstract

  Spliceosome-mediated RNA trans-splicing (SMaRT) provides an effective means to reprogram mRNAs and the proteins they encode. SMaRT technology has a broad range of applications, including RNA repair and molecular imaging, each governed by the nature of the sequences delivered by the pre-trans-splicing molecule. Here, we show the ability of SMaRT to optically image the expression of an exogenous gene at the level of pre-mRNA splicing in cells and living animals. Because of the modular design of pre-trans-splicing molecules, there is great potential to employ SMaRT to image the expression of any arbitrary gene of interest in living subjects. In this report, we describe a model system that demonstrates the feasibility of imaging gene expression by transsplicing in small animals. This represents a previously undescribed approach to molecular imaging of mRNA levels in living subjects.

  View details for DOI 10.1073/pnas.0402772101

  View details for Web of Science ID 000222037000037

  View details for PubMedID 15161977

  View details for PubMedCentralID PMC423257

• Spontaneous and controllable activation of suicide gene expression driven by the stress-inducible Grp78 promoter resulting in eradication of sizable human tumors HUMAN GENE THERAPY DONG, D. Z., Dubeau, L., Bading, J., Nguyen, K., Luna, M., Yu, H., Gazit-Bornstein, G., Gordon, E. M., Gomer, C., HALL, F. L., Gambhir, S. S., Lee, A. S. 2004; 15 (6): 553-561

  Abstract

  GRP78 is a stress-inducible chaperone protein with antiapoptotic properties that is overexpressed in transformed cells and cells under glucose starvation, acidosis, and hypoxic conditions that persist in poorly vascularized tumors. Previously we demonstrated that the Grp78 promoter is able to eradicate tumors using murine cells in immunocompetent models by driving expression of the HSV-tk suicide gene. Here, through the use of positron emission tomography (PET) imaging, we provide direct evidence of spontaneous in vivo activation of the HSV-tk suicide gene driven by the Grp78 promoter in growing tumors and its activation by photodynamic therapy (PDT) in a controlled manner. In this report, we evaluated whether this promoter can be applied to human cancer therapy. We observed that the Grp78 promoter, in the context of a retroviral vector, was highly activated by stress and PDT in three different types of human breast carcinomas independent of estrogen receptor and p53. Complete regression of sizable human tumors was observed after prodrug ganciclovir treatment of the xenografts in immunodeficient mice. In addition, the Grp78 promoter-driven suicide gene is strongly expressed in a variety of human tumors, including human osteosarcoma. In contrast, the activity of the murine leukemia virus (MuLV) long-terminal repeat (LTR) promoter varied greatly in different human breast carcinoma cell lines, and in some cases, stress resulted in partial suppression of the LTR promoter activity. In transgenic mouse models, the Grp78 promoter-driven transgene is largely quiescent in major adult organs but highly active in cancer cells and cancer-associated macrophages, which can diffuse to tumor necrotic sites devoid of vascular supply and facilitate cell-based therapy. Thus, transcriptional control through the use of the Grp78 promoter offers multiple novel approaches for human cancer gene therapy.

  View details for Web of Science ID 000222258500003

  View details for PubMedID 15212714

• Optical imaging of Renilla luciferase, synthetic Renilla luciferase, and firefly luciferase reporter gene expression in living mice JOURNAL OF BIOMEDICAL OPTICS Bhaumik, S., Lewis, X. Z., Gambhir, S. S. 2004; 9 (3): 578-586

  Abstract

  We have recently demonstrated that Renilla luciferase (Rluc) is a promising bioluminescence reporter gene that can be used for noninvasive optical imaging of reporter gene expression in living mice, with the aid of a cooled charged couple device (CCD) camera. In the current study, we explore the expression of a novel synthetic Renilla luciferase reporter gene (hRluc) in living mice, which has previously been reported to be a more sensitive reporter than native Rluc in mammalian cells. We explore the strategies of simultaneous imaging of both Renilla luciferase enzyme (RL) and synthetic Renilla luciferase enzyme (hRL):coelenterazine (substrate for RL/hRL) in the same living mouse. We also demonstrate that hRL:coelenterazine can yield a higher signal when compared to Firefly luciferase enzyme (FL): D-Luciferin, both in cell culture studies and when imaged from cells at the surface and from lungs of living mice. These studies demonstrate that hRluc should be a useful primary reporter gene with high sensitivity when used alone or in conjunction with other bioluminescence reporter genes for imaging in living rodents.

  View details for Web of Science ID 000221304400019

  View details for PubMedID 15189096

• Molecular imaging applications for immunology CLINICAL IMMUNOLOGY Hildebrandt, I. J., Gambhir, S. S. 2004; 111 (2): 210-224

  Abstract

  The use of multimodality molecular imaging has recently facilitated the study of molecular and cellular events in living subjects in a noninvasive and repetitive manner to improve the diagnostic capability of traditional assays. The noninvasive imaging modalities utilized for both small animal and human imaging include positron emission tomography (PET), single photon emission computed tomography (SPECT), magnetic resonance imaging (MRI), ultrasound, and computed tomography (CT). Techniques specific to small-animal imaging include bioluminescent imaging (BIm) and fluorescent imaging (FIm). Molecular imaging permits the study of events within cells, the examination of cell trafficking patterns that relate to inflammatory diseases and metastases, and the ability to rapidly screen new drug treatments for distribution and effectiveness. In this paper, we will review the current field of molecular imaging assays (especially those utilizing PET and BIm modalities) and examine how they might impact animal models and human disease in the field of clinical immunology.

  View details for Web of Science ID 000221681800008

  View details for PubMedID 15137954

• Quantitation of cell number by a positron emission tomography reporter gene strategy MOLECULAR IMAGING AND BIOLOGY Su, H., Forbes, A., Gambhir, S. S., Braun, J. 2004; 6 (3): 139-148

  Abstract

  An important potential of positron emission tomography (PET) is the capacity for quantitation of cell signals in an anatomic regions of interest. However, little is known about the constraints and parameters for using PET signal detection to establish cell numbers in regions of interest. In this study, we determined the correlation of PET signal to cell number, and characterized the cellular limit of detection for PET imaging.Cells expressing the herpes simplex virus type I thymidine kinase PET reporter gene (HSV1-sr39TK) were detected following accumulation of [(18)F]FHBG (9-[4-[(18)F]-fluoro-3-(hydroxymethyl) butyl]guanine) by microPET scanning and quantitation.When cells were cultured with [(18)F]FHBG in vitro, and then transferred to a model vascularized site, 73% retention was observed one hour post-transfer. Using this information, and the measured attenuation of PET signal in whole mouse scans, we assessed the per-cell uptake of [(18)F]FHBG in the vascularized site following standard parenteral administration of the substrate. We observed a cell number-dependent signal, with a limit of detection calculated as 10(6) cells in a region of interest of 0.1 mL volume. Quantitatively similar parameters were observed with stably tranfected N2a glioma cells and retrovirally transduced primary T lymphocytes.These methods and findings provide a strategy for quantitation of cellularity using PET imaging that has implications for both experimental models and clinical diagnosis.

  View details for DOI 10.1016/j.mibio.2004.02.001

  View details for Web of Science ID 000223467100005

  View details for PubMedID 15193248

• Molecular imaging of homodimeric protein-protein interactions in living subjects FASEB JOURNAL Massoud, T. F., Paulmurugan, R., Gambhir, S. S. 2004; 18 (7): 1105-?

  View details for DOI 10.1096/fj.03-1128fje

  View details for Web of Science ID 000221883200025

• MicroPET imaging of Cre-loxP-mediated conditional activation of a herpes simplex virus type 1 thymidine kinase reporter gene GENE THERAPY Sundaresan, G., Paulmurugan, R., Berger, F., Stiles, B., Nagayama, Y., Wu, H., Gambhir, S. S. 2004; 11 (7): 609-618

  Abstract

  Site-specific recombination tools such as the Cre-loxP system are used to create animal models where conditional gene deletion/activation studies are required. In the current proof of principle study, we have demonstrated that a PET reporter gene (PRG), the herpes simplex virus type 1 thymidine kinase (HSV1-tk), can be made to remain silent and can be activated by Cre-loxP-mediated recombination in cell culture and in living mice. An adenovirus carrying a silent HSV1-tk was tail-vein injected (1 x 10(9) PFU) in six transgenic mice that express Cre recombinase in their liver (Cre+) and in four control mice (Cre-). The liver-specific expression of the PRG in Cre+ mice was detected in the microPET following injection of the reporter probe, 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine ([(18)F]-FHBG). The [(18)F]-FHBG accumulation in the liver in terms of percent-injected dose per gram of tissue was 7.72+/-1.13 for the Cre+ mice and 0.10+/-0.02 for the Cre- mice (P<0.05) 48 h after adenoviral injection. These results were further validated by quantitative RT-PCR, western blotting and by in vitro assays for herpes simplex virus type 1 thymidine kinase enzyme activity. Thus by using the Cre-loxP system it is possible to modulate a PRG and noninvasively monitor the extent of Cre-loxP-mediated gene activation by imaging in a microPET scanner.

  View details for DOI 10.1038/sj.gt.3302194

  View details for Web of Science ID 000220281300006

  View details for PubMedID 14724687

• Tetraphenylphosphonium as a novel molecular probe for Imaging tumors JOURNAL OF NUCLEAR MEDICINE Min, J. J., Biswal, S., Deroose, C., Gambhir, S. S. 2004; 45 (4): 636-643

  Abstract

  Mitochondrial membrane potential (DeltaPsim)-dependent enhanced uptake of phosphonium salts, including (3)H-tetraphenylphosphonium ((3)H-TPP), in tumor cells, suggests the potential use of phosphonium salts as tracers for tumor imaging. In this study, we characterize the tumor accumulation of (3)H-TPP and compare it with (18)F-FDG in cell culture and in xenograft, metastatic, and inflammation models in living animals.(3)H-TPP and (3)H-FDG accumulation was compared in cell culture with a variety of cell lines in different glucose concentrations. Normal biodistribution and tumor uptake were assessed using nude mice with or without subcutaneous xenograft tumors (C6). To compare the accumulation of (3)H-TPP and (18)F-FDG in a metastatic tumor, severe combined immunodeficiency mice were tail-vein injected with human melanoma cell lines (A375-FL). To characterize the accumulation of (3)H-TPP and (18)F-FDG in inflammation, an inflammatory reaction was induced by subcutaneous injection of Complete Freund's Adjuvant in the left hind paw of Sprague-Dawley rat.The DeltaPsim data from a separate study and the current (3)H-TPP uptake data showed good correlation (r(2) = 0.82, P < 0.05). (3)H-TPP accumulation was significantly greater than that of (3)H-FDG for glucose >/=100 mg/dL. The biodistribution study of (3)H-TPP showed low uptake in most tissues but high accumulation in the heart and kidneys. (3)H-TPP accumulation in xenograft or metastatic tumors was comparable with that of (18)F-FDG, whereas (3)H-TPP accumulation in inflammatory tissues was markedly lower than that of (18)F-FDG.The sensitive tumor accumulation of (3)H-TPP with less propensity for inflammatory regions warrants further investigation of radiolabeled phosphonium analogs for tumor imaging in living subjects.

  View details for Web of Science ID 000220729700023

  View details for PubMedID 15073261

• Micro-positron emission tomography imaging of cardiac gene expression in rats using bicistronic adenoviral vector-mediated gene delivery CIRCULATION Chen, I. Y., Wu, J. C., Min, J. J., Sundaresan, G., Lewis, X., Liang, Q. W., Herschman, H. R., Gambhir, S. S. 2004; 109 (11): 1415-1420

  Abstract

  We have previously validated the use of micro-positron emission tomography (microPET) for monitoring the expression of a single PET reporter gene in rat myocardium. We now report the use of a bicistronic adenoviral vector (Ad-CMV-D2R80a-IRES-HSV1-sr39tk) for linking the expression of 2 PET reporter genes, a mutant rat dopamine type 2 receptor (D2R80a) and a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk), with the aid of an internal ribosomal entry site (IRES).Rat H9c2 cardiomyoblasts transduced with increasing titers of Ad-CMV-D2R80a-IRES-HSV1-sr39tk (0 to 2.5x10(8) pfu) were assayed 48 hours later for reporter protein activities, which were found to correlate well with viral titer (r2=0.96, P<0.001 for D2R80A; r2=0.98, P<0.001 for HSV1-sr39TK) and each other (r2=0.97; P<0.001). Experimental (n=8) and control (n=6) athymic rats underwent intramyocardial injection of up to 2x10(9) pfu of Ad-CMV-D2R80a-IRES-HSV1-sr39tk and saline, respectively. Forty-eight hours later and weekly thereafter, rats were assessed for D2R80a-dependent myocardial accumulation of 3-(2-[18F]fluoroethyl)spiperone ([18F]-FESP) and HSV1-sr39tk-dependent sequestration of 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]-FHBG) using microPET. Longitudinal [18F]-FESP and [18F]-FHBG imaging of experimental rats revealed a good correlation between the cardiac expressions of the 2 PET reporter genes (r2=0.73; P<0.001). The location of adenovirus-mediated transgene expression, as inferred from microPET images, was confirmed by ex vivo gamma counting of explanted heart.The IRES-based bicistronic adenoviral vector can potentially be used in conjunction with PET for indirect imaging of therapeutic gene expression by replacing 1 of the 2 PET reporter genes with a therapeutic gene of choice.

  View details for DOI 10.1161/01.CIR.0000121727.59564.5B

  View details for Web of Science ID 000220364700015

  View details for PubMedID 15007006

• Molecular imaging of drug-modulated protein-protein interactions in living subjects CANCER RESEARCH Paulmurugan, R., Massoud, T. F., Huang, J., Gambhir, S. S. 2004; 64 (6): 2113-2119

  Abstract

  Networks of protein interactions mediate cellular responses to environmental stimuli and direct the execution of many different cellular functional pathways. Small molecules synthesized within cells or recruited from the external environment mediate many protein interactions. The study of small molecule-mediated interactions of proteins is important to understand abnormal signal transduction pathways in cancer and in drug development and validation. In this study, we used split synthetic renilla luciferase (hRLUC) protein fragment-assisted complementation to evaluate heterodimerization of the human proteins FRB and FKBP12 mediated by the small molecule rapamycin. The concentration of rapamycin required for efficient dimerization and that of its competitive binder ascomycin required for dimerization inhibition were studied in cell lines. The system was dually modulated in cell culture at the transcription level, by controlling nuclear factor kappaB promoter/enhancer elements using tumor necrosis factor alpha, and at the interaction level, by controlling the concentration of the dimerizer rapamycin. The rapamycin-mediated dimerization of FRB and FKBP12 also was studied in living mice by locating, quantifying, and timing the hRLUC complementation-based bioluminescence imaging signal using a cooled charged coupled device camera. This split reporter system can be used to efficiently screen small molecule drugs that modulate protein-protein interactions and also to assess drugs in living animals. Both are essential steps in the preclinical evaluation of candidate pharmaceutical agents targeting protein-protein interactions, including signaling pathways in cancer cells.

  View details for PubMedID 15026351

• Bioluminescent monitoring of islet graft survival after transplantation MOLECULAR THERAPY Lu, Y. X., Dang, H., Middleton, B., Zhang, Z., Washburn, L., Campbell-Thompson, M., Atkinson, M. A., Gambhir, S. S., Tian, J., Kaufman, D. L. 2004; 9 (3): 428-435

  Abstract

  Islet transplantation offers a potential therapy to restore glucose homeostasis in type 1 diabetes patients. A method to image transplanted islets noninvasively and repeatedly would greatly assist studies of islet transplantation. Using recombinant adenovirus, we show that isolated rodent and human islets can be genetically engineered to express luciferase and then imaged after implantation into NOD-scid mice using a cooled charge-coupled device. The magnitude of the signal was dependent on the islet dose. Adenovirus-directed luciferase expression, however, rapidly attenuated. We next tested lentivirus vectors that should direct the long-term expression of reporter genes in transduced islets. Transplanted lentivirus-transduced islets restored euglycemia long term in streptozotocin-treated NOD-scid mice. The signal from implanted lentivirus-transduced islets was related directly to the implanted islet mass, and the signal did not attenuate over the observation period. Viral transduction, luciferase expression, and repeated imaging had no apparent long-term deleterious effects on islet function after implantation. These data demonstrate that the introduction of reporter genes into an isolated tissue allows the long-term monitoring of its survival following implantation. Such imaging technologies may allow earlier detection of graft rejection and the adjustment of therapies to prolong graft survival posttransplantation.

  View details for DOI 10.1016/j.mythe.2004.01.008

  View details for Web of Science ID 000220170100019

  View details for PubMedID 15006610

• Imaging tri-fusion multimodality reporter gene expression in living subjects CANCER RESEARCH Ray, P., De, A., Min, J. J., Tsien, R. Y., Gambhir, S. S. 2004; 64 (4): 1323-1330

  Abstract

  Imaging reporter gene expression in living subjects with various imaging modalities is a rapidly accelerating area of research. Applications of these technologies to cancer research, gene therapy, and transgenic models are rapidly expanding. We report construction and testing of several triple fusion reporter genes compatible with bioluminescence, fluorescence and positron emission tomography (PET) imaging. A triple fusion reporter vector harboring a bioluminescence synthetic Renilla luciferase (hrl) reporter gene, a reporter gene encoding the monomeric red fluorescence protein (mrfp1), and a mutant herpes simplex virus type 1 sr39 thymidine kinase [HSV1-truncated sr39tk (ttk); a PET reporter gene] was found to preserve the most activity for each protein component and was therefore investigated in detail. After validating the activities of all three proteins encoded by the fusion gene in cell culture, we imaged living mice bearing 293T cells transiently expressing the hrl-mrfp-ttk vector by microPET and using a highly sensitive cooled charge-coupled device camera compatible with both bioluminescence and fluorescence imaging. A lentiviral vector carrying the triple fusion reporter gene was constructed and used to isolate stable expressers by fluorescence-activated cell sorting. These stable 293T cells were further used to show good correlation (R(2) approximately 0.74-0.85) of signal from each component by imaging tumor xenografts in living mice with all three modalities. Furthermore, metastases of a human melanoma cell line (A375M) stably expressing the triple fusion were imaged by microPET and optical technologies over a 40-50-day time period in living mice. Imaging of reporter gene expression from single cells to living animals with the help of a single tri-fusion reporter gene will have the potential to accelerate translational cancer research.

  View details for Web of Science ID 000189245200019

  View details for PubMedID 14973078

• Postnatal hypoxic-ischemic brain injury alters mechanisms mediating neuronal glucose transport AMERICAN JOURNAL OF PHYSIOLOGY-REGULATORY INTEGRATIVE AND COMPARATIVE PHYSIOLOGY Zovein, A., Flowers-Ziegler, J., Thamotharan, S., Shin, D., Sankar, R., Nguyen, K., Gambhir, S., Devaskar, S. U. 2004; 286 (2): R273-R282

  Abstract

  We examined the effect of hypoxic ischemia and hypoxia vs. normoxia on postnatal murine brain substrate transporter concentrations and function. We detected a transient increase in the neuronal brain glucose transporter isoform (GLUT-3) in response to hypoxic ischemia after 4 h of reoxygenation. This increase was associated with no change in GLUT-1 (blood-brain barrier/glial isoform), monocarboxylate transporter isoforms 1 and 2, synapsin I (neuronal marker), or Bax (proapoptotic protein) but with a modest increase in Bcl-2 (antiapoptotic mitochondrial protein) protein concentrations. At 24 h of reoxygenation, the increase in GLUT-3 disappeared but was associated with a decline in Bcl-2 protein concentrations and the Bcl2:Bax ratio, an increase in caspase-3 enzyme activity (apoptotic effector enzyme), and extensive DNA fragmentation, which persisted later in time (48 h) only in the hippocampus. Hypoxia alone in the absence of ischemia was associated with a transient but modest increase in GLUT-3 and synapsin I protein concentrations, which did not cause significant apoptosis and/or necrosis. Assessment of glucose transporter function by 2-deoxyglucose (2-DG) uptake using two distinct techniques, namely positron emission tomography (PET) and the modified Sokoloff method, revealed a discrepancy due to glucose uptake by extracranial Harderian glands that masked the accurate detection of intracranial brain glucose uptake by PET scanning. The modified Sokoloff method assessing 2-DG uptake revealed that the transient increase in GLUT-3 was critical in protecting against a decline in brain glucose uptake. We conclude that hypoxic-ischemic brain injury is associated with transient compensatory changes targeted at protecting glucose delivery to fuel cellular energy metabolism, which then may delay the processes of apoptosis and cell necrosis.

  View details for DOI 10.1152/ajpregu.00160.2003

  View details for Web of Science ID 000187791500007

  View details for PubMedID 14525722

• Covalent disulfide-linked anti-CEA diabody allows site-specific conjugation and radiolabeling for tumor targeting applications PROTEIN ENGINEERING DESIGN & SELECTION Olafsen, T., Cheung, C. W., Yazaki, P. J., Li, L., Sundaresan, G., Gambhir, S. S., Sherman, M. A., Williams, L. E., Shively, J. E., Raubitschek, A. A., Wul, A. M. 2004; 17 (1): 21-27

  Abstract

  An engineered anti-carcinoembryonic antigen (CEA) diabody (scFv dimer, 55 kDa) was previously constructed from the murine anti-CEA T84.66 antibody. Tumor targeting, imaging and biodistribution studies in nude mice bearing LS174T xenografts with radiolabeled anti-CEA diabody demonstrated rapid tumor uptake and fast blood clearance, which are favorable properties for an imaging agent. Current radiolabeling approaches result in random modification of the protein surface, which may impair immunoreactivity especially for smaller antibody fragments. Site-specific conjugation approaches can direct modifications to reactive groups located away from the binding site. Here, cysteine residues were introduced into the anti-CEA diabody at three different locations, to provide specific thiol groups for chemical modification. One version (with a C-terminal Gly-Gly-Cys) existed exclusively as a disulfide-bonded dimer. This cysteine-modified diabody (Cys-diabody) retained high binding to CEA and demonstrated tumor targeting and biodistribution properties identical to the non-covalent diabody. Furthermore, following reduction of the disulfide bond, the Cys-diabody could be chemically modified using a thiol-specific bifunctional chelating agent, for radiometal labeling. Thus, the Cys-diabody provides a covalently linked alternative to conventional diabodies, which can be reduced and modified site-specifically. This format will provide a versatile platform for targeting a variety of agents to CEA-positive tumors.

  View details for DOI 10.1093/protein/gzh009

  View details for Web of Science ID 000220905100003

  View details for PubMedID 14985534

• Molecular Imaging Fundamentals Nuclear Medicine in Clinical Diagnosis & Treatment Gambhir, S. S., Massoud, T. edited by Gambhir, S. S., Ell, P. Churchill Livingstone Publishers. 2004; 3: 1845–1870
• Cell Biology Fundamentals Nuclear Medicine in Clinical Diagnosis & Treatment Gambhir, S. S., Deroose, C., Wall, Z. edited by Gambhir, S. S., Ell, P. Churchill Livingstone Publishers. 2004; 3: 1687–1712
• Nuclear Medicine in Clinical Diagnosis and Treatment Ell, P. J., Gambhir, S. S. Churchill Livingstone. 2004
• PET as a Tool in Multimodality Imaging of Gene Expression and Therapy Positron Emission Tomography: Basic Sciences De, A., Gambhir, S. S. edited by Bailey, D., Townsend, D., Valk, P., Maisey, M. Springer-Verlag: London. 2004: 343–367
• Decision Analysis Fundamentals Nuclear Medicine in Clinical Diagnosis & Treatment Gambhir, S. S., Schwimmer, J. edited by Gambhir, S. S., Ell, P. Churchill Livingstone Publishers. 2004; 3: 1911–1924
• Quantitative Assay Development for PET Positron Emission Tomography Gambhir, S. S. edited by Phelps, M. E. Raven Press. 2004: 125–216
• Brain PET Atlas Nuclear Medicine in Clinical Diagnosis & Treatment Silverman, D., Strommer, J., Marseille, D., Gambhir, S. S. edited by Gambhir, S. S., Ell, P. Churchill Livingstone Publishers. 2004; 3: 1475–1493
• PET of Cardiac Transgene Expression: Comparison of 2 Approaches Based on Herpesviral Thymidine Kinase Reporter Gene PET of Cardiac Transgene Expression: Comparison of 2 Approaches Based on Herpesviral Thymidine Kinase Reporter Gene Miyagawa, M. 2004; 45 (11): 1917-1923
• Gene therapy progress and prospects: Noninvasive imaging of gene therapy in living subjects GENE THERAPY Min, J. J., Gambhir, S. S. 2004; 11 (2): 115-125

  Abstract

  Recent progress in the development of noninvasive imaging technologies should allow molecular imaging to play a major role in the field of gene therapy. These tools have recently been validated in gene therapy models for continuous quantitative monitoring of the location(s), magnitude, and time variation of gene delivery and/or expression. This article reviews the use of radionuclide, magnetic resonance, and optical imaging technologies, as they have been used in imaging gene delivery and gene expression for gene therapy applications. The studies published to date lend support that noninvasive imaging tools will help to accelerate preclinical model validation, as well as allow for clinical monitoring of human gene therapy.

  View details for DOI 10.1038/sj.gt.3302191

  View details for Web of Science ID 000187909100001

  View details for PubMedID 14712295

• I-124-labeled engineered Anti-CEA minibodies and diabodies allow high-contrast, antigen-specific small-animal PET imaging of xenografts in athymic mice JOURNAL OF NUCLEAR MEDICINE Sundaresan, G., Yazaki, P. J., Shively, J. E., Finn, R. D., Larson, S. M., Raubitschek, A. A., Williams, L. E., Chatziioannou, A. F., Gambhir, S. S., Wu, A. M. 2003; 44 (12): 1962-1969

  Abstract

  Prolonged clearance kinetics have hampered the development of intact antibodies as imaging agents, despite their ability to effectively deliver radionuclides to tumor targets in vivo. Genetically engineered antibody fragments display rapid, high-level tumor uptake coupled with rapid clearance from the circulation in the athymic mouse/LS174T xenograft model. The anticarcinoembryonic antigen (CEA) T84.66 minibody (single-chain Fv fragment [scFv]-C(H)3 dimer, 80 kDa) and T84.66 diabody (noncovalent dimer of scFv, 55 kDa) exhibit pharmacokinetics favorable for radioimmunoimaging. The present work evaluated the minibody or diabody labeled with (124)I, for imaging tumor-bearing mice using a high-resolution small-animal PET system.Labeling was conducted with 0.2-0.3 mg of protein and 65-98 MBq (1.7-2.6 mCi) of (124)I using an iodination reagent. Radiolabeling efficiencies ranged from 33% to 88%, and immunoreactivity was 42% (diabody) or >90% (minibody). In vivo distribution was evaluated in athymic mice bearing paired LS174T human colon carcinoma (CEA-positive) and C6 rat glioma (CEA-negative) xenografts. Mice were injected via the tail vein with 1.9-3.1 MBq (53-85 microCi) of (124)I-minibody or with 3.1 MBq (85 microCi) of (124)I-diabody and imaged at 4 and 18 h by PET. Some mice were also imaged using (18)F-FDG 2 d before imaging with (124)I-minibody.PET images using (124)I-labeled minibody or diabody showed specific localization to the CEA-positive xenografts and relatively low activity elsewhere in the mice, particularly by 18 h. Target-to-background ratios for the LS174T tumors versus soft tissues using (124)I-minibody were 3.05 at 4 h and 11.03 at 18 h. Similar values were obtained for the (124)I-diabody (3.95 at 4 h and 10.93 at 18 h). These results were confirmed by direct counting of tissues after the final imaging. Marked reduction of normal tissue activity, especially in the abdominal region, resulted in high-contrast images at 18 h for the (124)I-anti-CEA diabody. CEA-positive tumors as small as 11 mg (<3 mm in diameter) could be imaged, and (124)I-anti-CEA minibodies, compared with (18)F-FDG, demonstrated highly specific localization.(124)I labeling of engineered antibody fragments provides a promising new class of tumor-specific probes for PET imaging of tumors and metastases.

  View details for Web of Science ID 000187219400024

  View details for PubMedID 14660722

• Synthesis of a new heterobifunctional linker, N-[4-(Aminooxy)butyl]maleimide, for facile access to a thiol-reactive F-18-Labeling agent BIOCONJUGATE CHEMISTRY Toyokuni, T., Walsh, J. C., Dominguez, A., Phelps, M. E., Barrio, J. R., Gambhir, S. S., Satyamurthy, N. 2003; 14 (6): 1253-1259

  Abstract

  A new heterobifunctional linker containing an aldehyde-reactive aminooxy group and a thiol-reactive maleimide group, namely N-[4-(aminooxy)butyl]maleimide, was synthesized as a stable HCl salt by O-alkylation of either N-hydroxyphthalimide or N-(4-monomethoxytrityl)hydroxylamine, followed by N-alkylation of maleimide, in an overall yield of 18% (seven steps) or 29% (five steps), respectively. This heterobifunctional linker allowed a simple and efficient synthesis of a maleimide-containing thiol-reactive (18)F-labeling agent. Thus, N-[4-[(4-[(18)F]fluorobenzylidene)aminooxy]butyl]maleimide (specific activity: approximately 3000 Ci/mmol at end of synthesis) was synthesized in two steps involving the preparation of 4-[(18)F]fluorobenzaldehyde, followed by its aminooxy-aldehyde coupling reaction to the heterobifunctional linker, with an overall radiochemical yield of approximately 35% (decay corrected) within approximately 60 min from end of bombardment. Initial (18)F-labeling experiments were carried out using a thiol-containing tripeptide glutathione (GSH) and a 5'-thiol-functionalized oligodeoxynucleotide (5'-S-ODN) in phosphate-buffered saline (PBS, pH 7.5). After standing at room temperature for 10 min, the (18)F-labeled GSH and 5'-S-ODN were obtained in (18)F-labeling yields of approximately 70% and approximately 5% (decay-corrected), respectively. The heterobifunctional linker is easy to synthesize and provides a facile access to the maleimide-containing thiol-reactive (18)F-labeling agent, which could be advantageously employed in the development of (18)F-labeled biomomolecules for use with positron emission tomography.

  View details for DOI 10.1021/bc034107y

  View details for Web of Science ID 000186725700025

  View details for PubMedID 14624642

• Optimization of adenoviral vectors to direct highly amplified prostate-specific expression for imaging and gene therapy MOLECULAR THERAPY Sato, M., Johnson, M., Zhang, L. Q., Zhang, B. H., Le, K., Gambhir, S. S., Carey, M., Wu, L. 2003; 8 (5): 726-737

  Abstract

  Gene expression-based imaging coupled to gene therapy will permit the prediction of therapeutic outcome. A significant challenge for successful gene therapy is to achieve a high-level of specific gene expression; however, tissue-specific promoters are weak. We postulate that if the weak activity of tissue-specific promoters can be amplified to the levels of strong viral promoters, which have been successful in preclinical scenarios, while retaining specificity, the therapeutic index of gene therapy can be greatly augmented. With this in mind, we developed a two-step transcriptional activation (TSTA) system. In this two-tiered system, a modified prostate-specific antigen promoter was employed to drive a potent synthetic transcriptional activator, GAL4-VP2. This, in turn, activated the expression of a GAL4-dependent reporter or therapeutic gene. Here we demonstrate that recombinant adenoviral vectors (Ads) in which we have incorporated prostate-targeted TSTA expression cassettes retain cell specificity and androgen responsiveness in cell culture and in animal models, as measured by noninvasive optical bioluminescence imaging. We investigated the mechanism of TSTA in different adenoviral configurations. In one configuration, both the activator and the reporter components are inserted into a single Ad (AdTSTA-FL). The activity of AdTSTA-FL exceeds that of a cytomegalovirus promoter-driven vector (AdCMV-FL), while maintaining tissue specificity. When the activator and reporter components are placed in two separate Ads, androgen induction is more robust than for the single AdTSTA-FL. Based on these findings, we hope to refine the TSTA Ads further to improve the efficacy and safety of prostate cancer gene therapy.

  View details for DOI 10.1016/j.ymthe.2003.08.016

  View details for Web of Science ID 000186597500010

  View details for PubMedID 14599805

• Comparison of [F-18]FHBG and [C-14]FIAU for imaging of HSV1-tk reporter gene expression: adenoviral infection vs stable transfection EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Min, J. J., Iyer, M., Gambhir, S. S. 2003; 30 (11): 1547-1560

  Abstract

  Earlier studies involving comparison of different reporter probes have shown conflicting results between pyrimidine nucleosides [e.g., 2'-fluoro-2'-deoxy-1-beta- d-arabinofuranosyl-5-iodouracil (FIAU)] and acycloguanosine derivatives [e.g., penciclovir (PCV), 9-(4-fluoro-3-hydroxymethylbutyl)guanine (FHBG)]. We hypothesized that this reported discrepancy may be related to how the reporter gene is delivered to the cells-stably transfected vs adenoviral infection. We directly compared the uptake characteristics of [(18)F]FHBG, [(3)H]PCV, and [(14)C]FIAU in cell culture and in vivo using an adenoviral mediated gene transfer model and stably transfected cells. We further compared the uptake of three reporter probes using both HSV1-tk and a mutant HSV1-sr39tk expressing cells to assess the optimal reporter probe/reporter gene combination. [(14)C]FIAU accumulation was greater than that of [(3)H]PCV and [(18)F]FHBG in control cells and in HSV1-tk stably transfected cells ( P<0.001). After infection of C6 cells with AdCMV- HSV1-tk (1.5x10(8) pfu), [(18)F]FHBG and [(3)H]PCV accumulation was significantly greater than that of [(14)C]FIAU ( P<0.01). [(18)F]FHBG and [(3)H]PCV accumulated to a significantly greater extent than [(14)C]FIAU in C6-stb-sr39tk+ and AdCMV- HSV1-sr39tk infected C6 cells ( P<0.001). Results from the nude mice supported the results in cell culture. [(14)C]FIAU led to significantly higher %ID/g in C6-stb-tk+ xenografts than [(18)F]FHBG ( P<0.05); however, compared with [(14)C]FIAU, [(18)F]FHBG led to as high %ID/g in HSV1-tk expressing hepatocytes and to significantly greater %ID/g in C6-stb-sr39tk+ xenografts and HSV1-sr39tk expressing hepatocytes. Dynamic sequential images showed that [(18)F]FHBG was well retained in HSV1-sr39tk expressing cells (C6-stb-sr39tk+) for at least 4 h after injection, while it was rapidly cleared from HSV1-tk expressing cells (MH3924A-stb-tk+). [(14)C]FIAU accumulated in HSV1-tk stably expressing cells to a greater extent than either [(3)H]PCV or [(18)F]FHBG. However, the accumulation of [(3)H]PCV and [(18)F]FHBG in adenoviral infected C6 cells or hepatocytes was equivalent to or greater than that of [(14)C]FIAU. These results may be due to intracellular biochemical changes (e.g., thymidine) when cells are infected with adenovirus. For adenoviral studies, the [(18)F]FHBG/ HSV1-sr39tk combination was shown to be more sensitive than the [(14)C]FIAU/ HSV1-tk combination HSV1-tk.

  View details for DOI 10.1007/s00259-003-1238-6

  View details for Web of Science ID 000186139700017

  View details for PubMedID 14579096

• Atorvastatin prevents RhoC isoprenylation, invasion, and metastasis in human melanoma cells MOLECULAR CANCER THERAPEUTICS Collisson, E. A., Kleer, C., Wu, M., De, A., Gambhir, S. S., Merajver, S. D., Kolodney, M. S. 2003; 2 (10): 941-948

  Abstract

  Melanoma is a deadly cancer due to its propensity to metastasize. Pharmacological inhibition of cell motility may benefit patients with cutaneous melanoma by preventing metastasis to internal organs. The Rho GTPases are signaling molecules that drive metastasis by controlling cell motility. We found RhoC to be expressed in clinical melanoma specimens and hypothesized that inhibiting its activation might prevent metastasis. Some Rho proteins, such as RhoC, depend on posttranslational geranylgeranylation for biological activity. We investigated the effect that Atorvastatin, a 3-hydroxy 3-methylglutaryl CoA (HMG-CoA) reductase inhibitor that prevents Rho geranylgeranylation, had on subcellular localization and activity of Rho proteins as well as the metastatic ability of melanoma cells. Atorvastatin inhibited Rho activation and reverted the metastatic phenotype of human melanoma cells in vitro. Moreover, Atorvastatin, at plasma levels comparable to those used to treat of hypercholesterolemia, inhibited in vivo metastasis of melanoma cells overexpressing RhoC. These results support further examination of statins for primary prophylaxis of melanoma metastasis.

  View details for Web of Science ID 000186054800002

  View details for PubMedID 14578459

• Molecular imaging of cardiac cell transplantation in living animals using optical bioluminescence and positron emission tomography CIRCULATION Wu, J. C., Chen, I. Y., Sundaresan, G., Min, J. J., De, A., Qiao, J. H., Fishbein, M. C., Gambhir, S. S. 2003; 108 (11): 1302-1305

  Abstract

  The current method of analyzing myocardial cell transplantation relies on postmortem histology. We sought to demonstrate the feasibility of monitoring transplanted cell survival in living animals using molecular imaging techniques.For optical bioluminescence charged-coupled device imaging, rats (n=20) underwent intramyocardial injection of embryonic rat H9c2 cardiomyoblasts (3x10(6) to 5x10(5)) expressing firefly luciferase (Fluc) reporter gene. Cardiac bioluminescence signals were present for more than 2 weeks with 3x10(6) cells: day 1 (627 000+/-15%), day 2 (346 100+/-21%), day 4 (112 800+/-20%), day 8 (78 860+/-24%), day 12 (67 780+/-12%), and day 16 (62 200+/-5% p x s(-1) x cm(2-1) x sr(-1)). For micro-positron emission tomography imaging, rats (n=20) received cardiomyoblasts (3x10(6)) expressing mutant herpes simplex type 1 thymidine kinase (HSV1-sr39tk) reporter gene. Detailed tomography of transplanted cells is shown by 9-(4-[18F]-fluoro-3hydroxymethylbutyl)guanine ([18F]-FHBG) reporter probe and nitrogen-13 ammonia ([13N]-NH3) perfusion images. Within the transplanted region, there was a 4.48+/-0.71-fold increase of in vivo [18F]-FHBG activity and a 4.01+/-0.51-fold increase of ex vivo gamma counting compared with control animals. Finally, the in vivo images of cell survival were confirmed by ex vivo autoradiography, histology, immunohistochemistry, and reporter protein assays.The location(s), magnitude, and survival duration of embryonic cardiomyoblasts were monitored noninvasively. With further development, molecular imaging studies should add critical insights into cardiac cell transplantation biology.

  View details for DOI 10.1161/01.CIR.0000091252.20010.6E

  View details for Web of Science ID 000185328800006

  View details for PubMedID 12963637

• Treatment of metastatic melanoma with an orally available inhibitor of the Ras-Raf-MAPK cascade CANCER RESEARCH Collisson, E. A., De, A., Suzuki, H., Gambhir, S. S., Kolodney, M. S. 2003; 63 (18): 5669-5673

  Abstract

  The Ras-Raf-MAPK pathway is constitutively activated in the majority of melanomas because of a mutation in the BRAF gene. It has been hypothesized that activation of this pathway is crucial for the genesis and maintenance of melanoma and therefore represents an attractive clinical target for metastatic disease. We synthesized a previously characterized MAP kinase kinase inhibitor to test the effect that blocking the Ras-Raf-MAPK pathway would have on the establishment and maintenance of melanoma metastases. Oral administration of CI 1040 inhibited formation of pulmonary metastases and caused rapid regression of established pulmonary metastases in the mouse. Our findings indicate that Ras-Raf-MAPK activation provides crucial signals for the survival of melanoma cells at ectopic sites and that the pharmacological inhibition of this pathway is a promising target for melanoma therapy.

  View details for Web of Science ID 000185672600003

  View details for PubMedID 14522881

• LABAZ1: A metastatic tumor model for renal cell carcinoma expressing the carbonic anhydrase type 9 tumor antigen CANCER RESEARCH Zisman, A., Pantuck, A. J., Bui, M. H., Said, J. W., Caliliw, R. R., Rao, N., Shintaku, P., Berger, F., Gambhir, S. S., Belldegrun, A. S. 2003; 63 (16): 4952-4959

  Abstract

  A metastatic renal cell carcinoma (RCC) tumor model xenograft that expresses the targetable, membrane-bound tumor-associated antigen carbonic anhydrase type 9 (CA IX) is described. The xenograft, established from a high-grade type-2 chromophil RCC (cRCC), has been serially transplanted in immune compromised mice, in which it grows orthotopically under the renal capsule, doubling its size every 9 weeks and sending metastases to the lung and liver at approximately 20 weeks. Tumors were capable of being imaged using a micro-PET (micro-positron emission tomograph) with an 18-fluorodeoxyglucose (18-FDG) tracer. Subsequent xenograft generations have conserved immunohistochemical and ultrastructural properties typical for malignant renal epithelium-derived neoplasia (vimentin+, CK-19+, CA IX+ with hypoxia-inducible factor (HIF)-1 alpha constitutive expression) and have demonstrated extensive proliferation, lack of apoptosis, severe genetic alterations, and molecular expression alterations; transforming growth factor beta 1 (TGF-beta 1), hepatocyte growth factor (HGF), proto-oncogene (c-met), matrix metalloproteinase (MMP)-1, and vascular endothelial growth factor (VEGF) C and D were overexpressed, whereas human epidermal growth factor receptor (HER)-2, MMP-2 and MMP-9, VEGF-R3, p53, and p27 were severely down-regulated, suggesting a proangiogenic environment, local invasiveness, and facilitated lymphatic metastasis. Altogether, LABAZ1 provides a relevant and flexible model to study the biology of cRCC, the role of CA IX in RCC tumorigenesis, progression, and metastasis, and a platform for testing new targeted therapeutic strategies.

  View details for Web of Science ID 000184948400032

  View details for PubMedID 12941820

• Interrogating androgen receptor function in recurrent prostate cancer CANCER RESEARCH Zhang, L. Q., Johnson, M., Le, K. H., Sato, M., Ilgan, R., Iyer, M., Gambhir, S. S., Wu, L., Carey, M. 2003; 63 (15): 4552-4560

  Abstract

  The early androgen-dependent (AD) phase of prostate cancer is dependent on the androgen receptor (AR). However, it is unclear whether AR is fully functional in recurrent prostate cancer after androgen withdrawal. To address this issue we interrogated AR signaling in AD and recurrent prostate cancer xenografts using molecular imaging, chromatin immunoprecipitation, and immunohistochemistry. In the imaging experiments, an adenovirus bearing a two-step transcriptional activation cassette, which amplifies AR-dependent firefly luciferase reporter gene activity, was injected into tumors implanted into severe combined immunodeficiency mice. A charge-coupled device optical imaging system detected the initial loss and then resumption of AR transcriptional activity in D-luciferin-injected mice as tumors transitioned from AD to recurrent growth. The results of chromatin immunoprecipitation and immunohistochemical localization experiments correlated with the Ad two-step transcriptional activation imaging signal. AR localized to the nucleus and bound to the endogenous prostate-specific antigen enhancer in AD tumors but exited the nucleus and dissociated from the enhancer upon castration. However, AR reentered the nucleus and rebound the prostate-specific antigen enhancer as the cancer transitioned into the recurrent phase. Surprisingly, RNA polymerase II and the general factor TFIIB remained bound to the gene throughout the transition. Our data support the concept that AR is fully functional in recurrent cancer and suggest a model by which a poised but largely inactive transcription complex facilitates reactivation by AR at castrate levels of ligand.

  View details for Web of Science ID 000184562500039

  View details for PubMedID 12907631

• AMIDE: a free software tool for multimodality medical image analysis. Molecular imaging Loening, A. M., Gambhir, S. S. 2003; 2 (3): 131-137

  Abstract

  Amide's a Medical Image Data Examiner (AMIDE) has been developed as a user-friendly, open-source software tool for displaying and analyzing multimodality volumetric medical images. Central to the package's abilities to simultaneously display multiple data sets (e.g., PET, CT, MRI) and regions of interest is the on-demand data reslicing implemented within the program. Data sets can be freely shifted, rotated, viewed, and analyzed with the program automatically handling interpolation as needed from the original data. Validation has been performed by comparing the output of AMIDE with that of several existing software packages. AMIDE runs on UNIX, Macintosh OS X, and Microsoft Windows platforms, and it is freely available with source code under the terms of the GNU General Public License.

  View details for PubMedID 14649056

• Optical imaging of transferrin targeted PEI/DNA complexes in living subjects GENE THERAPY Hildebrandt, I. J., Iyer, M., Wagner, E., Gambhir, S. S. 2003; 10 (9): 758-764

  Abstract

  Noninvasive optical bioluminescence imaging systems are important tools for evaluating gene expression in vivo for study of individual and temporal variation in a living animal. In this report, we demonstrate that expression of the firefly luciferase reporter gene (fl) delivered by transferrin (Tf) targeted polyethylenimine (PEI) complexes with, or without, poly(ethylene glycol) (PEG) modifications can be imaged in living A/J mice bearing N2A tumors using a cooled charged coupled device (CCD) camera. Tf-PEI-PEG, Tf-PEI, and PEI (positive control) complexes were tail-vein injected and mice were imaged at 5, 24, 48, and 72 h after complex injection. After imaging, the organs were analyzed ex vivo for firefly luciferase protein (FL) activity. The Tf and PEG modified formulations show significantly (P<0.05) higher FL activity in vivo and ex vivo at the tumor as compared to other organs, including the lungs (a site of high expression with PEI, the positive control). Furthermore, the in vivo bioluminescent signal correlated well (R(2)=0.83) with ex vivo FL activity. These data support that noninvasive imaging of fl reporter expression can be used to monitor the specificity of Tf-PEI and Tf-PEI-PEG polyplex targeting of N2A tumors in A/J mice.

  View details for DOI 10.1038/sj.gt.3301939

  View details for Web of Science ID 000182482200005

  View details for PubMedID 12704414

• Noninvasive imaging of lentiviral-mediated reporter gene expression in living mice MOLECULAR THERAPY De, A., Lewis, X. Z., Gambhir, S. S. 2003; 7 (5): 681-691

  Abstract

  Lentiviral-mediated gene delivery holds significant promise for sustained gene expression within living systems. Vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 1-based lentiviral vectors can be used to introduce transgenes in a broad spectrum of dividing as well as nondividing cells. In the current study, we construct a lentiviral vector carrying two reporter genes separated by an internal ribosomal entry site and utilize that virus in delivering both genes into neuroblastoma cells in cell culture and into cells implanted in living mice. We utilize two reporter genes, a mutant herpes simplex virus type 1 (HSV1) sr39tk as a reporter gene compatible with positron emission tomography (PET) and a bioluminescent optical reporter gene, firefly luciferase (Fluc), to image expression in living mice by an optical charge-coupled device (CCD) camera. By using this lentivirus, neuroblastoma (N2a) cells are stably transfected and a high correlation (R(2) = 0.91) between expressions of the two reporter genes in cell culture is established. Imaging of both reporter genes using microPET and optical CCD camera in living mice is feasible, with the optical approach being more sensitive, and a high correlation (R(2) = 0.86) between gene expressions is again observed in lentiviral-infected N2a tumor xenografts. Indirect imaging of HSV1-sr39tk suicide gene therapy utilizing Fluc is also feasible and can be detected with increased sensitivity by using the optical CCD. These preliminary results validate the use of lentiviral vectors carrying reporter genes for multimodality imaging of gene expression and should have many applications, including imaging of xenografts, metastasis, and cell trafficking as well as noninvasive monitoring of lentiviral-mediated gene delivery and expression.

  View details for DOI 10.1016/S1525-0016(03)00070-4

  View details for Web of Science ID 000182645800016

  View details for PubMedID 12718911

• Positron emission tomography in diagnosis and management of invasive breast cancer: current status and future perspectives. Clinical breast cancer Wu, D., Gambhir, S. S. 2003; 4: S55-63

  Abstract

  [18F]fluorodeoxyglucose positron emission tomography (FDG-PET) is a metabolic imaging modality that has increasing applications in oncology, neurology, and cardiology. Among the oncology applications, breast cancer is one of the most extensively studied diseases. FDG-PET has been performed for diagnosis, staging, and restaging of invasive breast cancer and for monitoring responsiveness to therapies. At the present time, the results of FDG-PET in detection of primary breast cancer and axillary staging are mixed and inconclusive. However, results demonstrating the superiority of FDG-PET over anatomic imaging modalities in detection of distant metastasis, recurrence, and monitoring therapies are relatively well documented. These applications have been accepted by medical professionals and the public, as evidenced by a recent decision by the Centers for Medicare and Medicaid Services (formerly Health Care Financing Agency) to provide coverage for the procedure. Future trends in this exciting area include development of novel breast cancer-specific PET radiopharmaceuticals and use of dedicated breast PET technologies for scans of breast/axillary lesions. PET/computed tomography technology, which combines anatomic and molecular/biochemical information, is also rapidly proliferating and should help to further improve the management of patients with breast cancer. The role of FDG-PET in breast cancer is increasing and evolving, and this metabolic imaging modality, in conjunction with newer tracers and other anatomic imaging methods, should improve diagnosis and management of patients with breast cancer

  View details for PubMedID 12756080

• Monitoring protein-protein interactions using split synthetic renilla luciferase protein-fragment-assisted complementation ANALYTICAL CHEMISTRY Paulmurugan, R., Gambhir, S. S. 2003; 75 (7): 1584-1589

  Abstract

  In this study we developed an inducible synthetic renilla luciferase protein-fragment-assisted complementation-based bioluminescence assay to quantitatively measure real time protein-protein interactions in mammalian cells. We identified suitable sites to generate fragments of N and C portions of the protein that yield significant recovered activity through complementation. We validate complementation-based activation of split synthetic renilla luciferase protein driven by the interaction of two strongly interacting proteins, MyoD and Id, in five different cell lines utilizing transient transfection studies. The expression level of the system was also modulated by tumor necrosis factor alpha through NFkappaB-promoter/enhancer elements used to drive expression of the N portion of synthetic renilla luciferase reporter gene. This new system should help in studying protein-protein interactions and when used with other split reporters (e.g., split firefly luciferase) should help to monitor different components of an intracellular network.

  View details for DOI 10.1021/ac020731c

  View details for Web of Science ID 000181993600013

  View details for PubMedID 12705589

• MicroPET imaging of prostate cancer in LNCAP-SR39TK-GFP mouse xenografts PROSTATE Yang, H. H., Berger, F., Tran, C., Gambhir, S. S., Sawyers, C. L. 2003; 55 (1): 39-47

  Abstract

  The aim of this study was to develop models that allow serial, noninvasive imaging of human prostate cancer cells in immunodeficient mice using a dedicated small animal positron emission tomography scanner (microPET).LNCaP tumor cells were stably transduced ex-vivo with the mutant herpes simplex virus type 1 thymidine kinase (HSV-sr39tk) PET reporter gene and green fluorescent protein (GFP). The stably transduced LNCaP cells were then enriched via fluorescent cell sorting and implanted into SCID mice. Beginning 2 weeks after tumor cell inoculation, mice were repeatedly scanned by microPET performed 1 hr after tail-vein injection of approximately 200 muCi Fluorine-18 labeled penciclovir ((18)F-FHBG). PET-images were correlated to tumor size, % injected dose (ID)/g tumor tissue, PSA levels, autoradiography, and histology.Monitoring LNCaP xenografts using microPET and our reporter gene approaches is feasible. MicroPET was capable of detecting subcutaneous tumors as small as 3 mm in diameter (approximately 0.2% ID/g). The magnitude of (18)F-FHBG-uptake in PET-images correlated with the tumor volumes and the serum PSA levels. Other non-HSV1-TK-specific tracers were also studied. While (18)F-flurodeoxyglucose ((18)F-FDG) gave poor imaging results in LNCaP cells, (11)C-acetate gave satisfactory images.We demonstrated the feasibility of monitoring prostate cancer xenografts in a mouse model using microPET and the HSV1-sr39tk PET reporter gene/(18)F-FHBG reporter probe system. Extension of this approach may allow repetitive imaging of tumor metastases.

  View details for DOI 10.1002/pros.10208

  View details for Web of Science ID 000181973300005

  View details for PubMedID 12640659

• Positron emission tomography imaging of cellular cardiomyoplasty. Wu, J. C., Chen, I. Y., De, A., Min, J. J., Sundaresan, G., Qiao, J. H., Fishbein, M. C., Gambhir, S. S. ELSEVIER SCIENCE INC. 2003: 446A

  View details for Web of Science ID 000181669501932

• Optical bioluminescence and positron emission tomography imaging of a novel fusion reporter gene in tumor xenografts of living mice CANCER RESEARCH Ray, P., Wu, A. M., Gambhir, S. S. 2003; 63 (6): 1160-1165

  Abstract

  Noninvasive imaging of reporter gene expression using various imaging modalitiesis playing an increasingly important role in defining molecular events in the field of cancer biology, cell biology, and gene therapy. In this study, a novel reporter vector was constructed encoding a fusion protein comprised of a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) (tk), a positron emission tomography (PET) reporter gene, and renilla luciferase (rl), a bioluminescence optical reporter gene joined by a 20 amino acid long spacer sequence. We validated the activity of the two enzymes encoded by the fusion protein (tk(20)rl) in cell culture. Then, tumors stably expressing the tk(20)rl fusion gene were imaged both by microPET and optically using a cooled charge coupled device camera in xenograft-bearing living mice. Using a single fusion reporter (PET/optical) gene should accelerate the validation of reporter gene approaches developed in cell culture for translation into preclinical and clinical models.

  View details for Web of Science ID 000181702300003

  View details for PubMedID 12649169

• Molecular imaging in living subjects: seeing fundamental biological processes in a new light GENES & DEVELOPMENT Massoud, T. F., Gambhir, S. S. 2003; 17 (5): 545-580

  View details for DOI 10.1101/gad.1047403

  View details for Web of Science ID 000181502300001

  View details for PubMedID 12629038

• Quantitative imaging of the T cell antitumor response by positron-emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Dubey, P., Su, H., Adonai, N., Du, S. Y., Rosato, A., Braun, J., Gambhir, S. S., Witte, O. N. 2003; 100 (3): 1232-1237

  Abstract

  We describe a noninvasive, quantitative, and tomographic method to visualize lymphocytes within the whole animal. We used positron-emission tomography (PET) to follow the localization of adoptively transferred immune T lymphocytes. Splenic T cells from animals that had rejected a Moloney murine sarcoma virus/Moloney murine leukemia virus (M-MSV/M-MuLV)-induced tumor were marked with a PET reporter gene, injected into tumor-bearing mice, and imaged in a microPET by using a substrate specific for the reporter. Specific localization of immune T cells to the antigen-positive tumor was detected over time, by sequential imaging of the same animals. Naive T cells did not localize to the tumor site, indicating that preimmunization was required. Autoradiography and immunohistochemistry analysis corroborated the microPET data. The method we have developed can be used to assess the effects of immunomodulatory agents intended to potentiate the immune response to cancer, and can also be useful for the study of other cell-mediated immune responses, including autoimmunity.

  View details for DOI 10.1073/pnas.0337418100

  View details for Web of Science ID 000180838100084

  View details for PubMedID 12547911

• Positron-emission tomography reporter gene expression imaging in rat myocardium CIRCULATION Inubushi, M., Wu, J. C., Gambhir, S. S., Sundaresan, G., Satyamurthy, N., Namavari, M., Yee, S., Barrio, J. R., Stout, D., Chatziioannou, A. F., Wu, L. L., Schelbert, H. R. 2003; 107 (2): 326-332

  Abstract

  This study examines the quantitative accuracy, detection sensitivity, and time course of imaging the expression of a mutant herpes simplex type-1 virus thymidine kinase (HSV1-sr39tk) PET reporter gene in rat myocardium by using the PET reporter probe 9-(4-[18F]-Fluoro-3-Hydroxymethylbutyl)-Guanine ([18F]-FHBG) and a small-animal PET (microPET).In 40 rats, adenovirus expressing HSV1-sr39tk driven by a cytomegalovirus promoter (Ad-CMV-HSV1-sr39tk, 1x10(6) to 1x10(9) pfu) was injected through a thoracotomy directly into the left ventricular myocardium. After 3 days, myocardial perfusion was imaged with [13N]-ammonia for delineating the left ventricular myocardium, followed by imaging the expression of the reporter gene with intravenous [18F]-FHBG. The total myocardial [18F]-FHBG accumulation was quantified in percent of injected dose (%ID). Immunohistochemistry and autoradiography demonstrated HSV1-sr39tk enzyme (HSV1-sr39TK) and accumulation of [18F]-FHBG in the inoculated myocardium in 3 rats each. In 24 rats with various viral titers, the %ID was correlated with ex vivo well counting (r2=0.981, P<0.0001) and myocardial HSV1-sr39TK activity by tissue enzyme activity assay (r2=0.790, P<0.0001). Myocardial [18F]-FHBG accumulation was identified at viral titers down to 1x10(7) pfu. In 6 rats serially imaged up to day 17, myocardial [18F]-FHBG accumulation on microPET peaked on days 3 to 5 and was no longer identified on days 10 to 17.HSV1-sr39tk reporter gene expression can be monitored with [18F]-FHBG and microPET in rat myocardium quantitatively and serially with high detection sensitivity. Cardiac PET reporter gene imaging offers the potential of monitoring the expression of therapeutic genes in cardiac gene therapy.

  View details for DOI 10.1161/01.CIR.0000044385.60972.AE

  View details for Web of Science ID 000180785700022

  View details for PubMedID 12538436

• Molecular Imaging with Positron Emission Tomography. Discussions in PET Imaging. Diagnostic Imaging Gambhir SS 2003: 1-4
• Gene and Cell Therapy Gene Therapy: Therapeutic Mechanisms and Strategies Biswal, S., Gambhir, S. S., Templeton, N. Marcel Dekker, Inc: New York . 2003: 447–480
• PET in Imaging Gene Expression and Therapy Positron Emission Tomography: Basic Science and Clinical Practice De, A., Gambhir, S. S. edited by Bailey, D., Townsend, D., Valk, P., Maisey, M. Springer-Verlag. 2003: 845–868
• Quantitation of the regional blood flow in the interventricular septum using positron emission tomography and nitrogen-13 ammonia EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Hove, J. D., Gambhir, S. S., Kofoed, K. F., Freiberg, J., Kelbaek, H. 2003; 30 (1): 109-116

  Abstract

  The purpose of the present study was to evaluate the effect of spillover of activity from the right ventricle (RV) on quantitation of the regional myocardial blood flow in the septum. Thirty-one healthy volunteers, 31 patients with ischemic heart disease, 7 patients with severe congestive heart failure, and 6 heart transplant patients underwent positron emission tomography (PET) with nitrogen-13 ammonia. Quantitation of the regional myocardial blood flow in the septum was performed using both a conventional two-compartment model and a previously validated two-compartment model taking RV spillover into account. Unaccounted RV spillover resulted in significant underestimation of the regional myocardial blood flow in the septum. The amount of underestimation was primarily dependent on the magnitude of spillover and the dispersion between the right and the left ventricular input functions. In healthy volunteers, the flow error was small but significant: on average 6% (range 5%-29%, P<0.00001), compared with 27% (range 0%-88%, P<0.002) in the group of patients with severe congestive heart failure, who had the most considerable amount of RV spillover. In the group of patients with ischemic heart disease and the group of heart transplant patients the flow errors were 10% (range 0%-55%, P<0.00001) and 6% (range 1%-19%, P<0.01), respectively. It is concluded that flow quantitation in the septum is significantly affected by RV spillover, resulting in a considerable underestimation of the septal blood flow unless correction is performed.

  View details for DOI 10.1007/s00259-002-1014-z

  View details for Web of Science ID 000180691800016

  View details for PubMedID 12483417

• 6th ASNC Invitational Conference on Nuclear Cardiology ? Panel on Gene Product Imaging. Journal of Nuclear Cardiology Bengel FM, Dewanjee M, Dichek D, Gambhir SS, Gelovani J, Zaret B. 2003; 10: 223-257
• Monitoring adenoviral DNA delivery, using a mutant herpes simplex virus type 1 thymidine kinase gene as a PET reporter gene GENE THERAPY Liang, Q., Nguyen, K., Satyamurthy, N., Barrio, J. R., Phelps, M. E., Gambhir, S. S., Herschman, H. R. 2002; 9 (24): 1659-1666

  Abstract

  Current gene therapy protocols often suffer from an inability to monitor the site, level and persistence of gene expression following somatic DNA delivery. Herpes simplex virus 1 thymidine kinase (HSV1-tk) is currently under intensive investigation as a reporter gene for in vivo imaging of reporter gene expression. The presence of the HSV1-tk reporter gene is repetitively and non-invasively monitored by systemic injection of positron-emitting, radionuclide-labeled thymidine analogues or acycloguanosine HSV1-TK substrates and subsequent detection, by positron emission tomography, of trapped, phosphorylated product. To improve the efficacy of the HSV1-tk PET reporter gene system, both alternative substrates and mutations in the HSV1-tk gene have been described. We used a replication defective adenovirus to deliver the HSV1-sr39tk mutant enzyme and the wild-type HSV1-tk enzyme to mice. HSV1-sr39TK demonstrates greater sensitivity than wild-type HSV1-TK enzyme in vivo, using 9-[(4-[(18)F]fluoro-3-hydroxymethylbutyl)guanine as probe, following adenovirus-mediated hepatic expression in mice. Using this adenoviral delivery system, the location, magnitude and duration of HSV1-sr39tk PET reporter gene expression could be non-invasively, quantitatively and repetitively monitored for over 3 months by microPET.

  View details for Web of Science ID 000179898900003

  View details for PubMedID 12457279

• Noninvasive imaging of protein-protein interactions in living subjects by using reporter protein complementation and reconstitution strategies PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Paulmurugan, R., Umezawa, Y., Gambhir, S. S. 2002; 99 (24): 15608-15613

  Abstract

  In this study we have developed bioluminescence-imaging strategies to noninvasively and quantitatively image protein-protein interactions in living mice by using a cooled charge-coupled device camera and split reporter technology. We validate both complementation and intein-mediated reconstitution of split firefly luciferase proteins driven by the interaction of two strongly interacting proteins, MyoD and Id. We use transient transfection of cells and image MyoD-Id interaction after induction of gene expression in cell culture and in cells implanted into living mice. Techniques to study protein-protein interactions in living subjects will allow the study of cellular networks, including signal transduction pathways, as well as development and optimization of pharmaceuticals for modulating protein-protein interactions.

  View details for DOI 10.1073/pnas.242594299

  View details for Web of Science ID 000179530000065

  View details for PubMedID 12438689

  View details for PubMedCentralID PMC137764

• Noninvasive imaging of cationic lipid-mediated delivery of optical and PET reporter genes in living mice MOLECULAR THERAPY Iyer, M., Berenji, M., Templeton, N. S., Gambhir, S. S. 2002; 6 (4): 555-562

  Abstract

  Gene therapy involves the safe and effective delivery of one or more genes of interest to target cells in vivo. The advantages of using nonviral delivery systems include ease of preparation, low toxicity, and weak immunogenicity. Nonviral delivery methods, when combined with a noninvasive, clinically applicable imaging assay, will greatly aid in the optimization of gene therapy approaches for cancer. We demonstrate cationic lipid-mediated noninvasive monitoring of reporter gene expression of firefly (Photinus pyralis) luciferase (fl) and a mutant herpes simplex virus type I thymidine kinase (HSV1-sr39tk, tk) in living mice using a cooled charge coupled device (CCD) camera and positron emission tomography (PET), respectively. We observe a high level of fl and tk reporter gene expression predominantly in the lungs after a single injection of the extruded DOTAP:cholesterol DNA liposome complexes by way of the tail vein, seen to be time- and dose-dependent. We observe a good correlation between the in vivo bioluminescent signal and the ex vivo firefly luciferase enzyme (FL) activity in different organs. We further demonstrate the feasibility of noninvasively imaging both optical and PET reporter gene expression in the same animal using the CCD camera and microPET, respectively.

  View details for DOI 10.1006/mthe.2002.0700

  View details for Web of Science ID 000178472200018

  View details for PubMedID 12385291

• Whole-body skeletal imaging in mice utilizing microPET: optimization of reproducibility and applications in animal models of bone disease EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Berger, F., Lee, Y. P., Loening, A. M., Chatziioannou, A., Freedland, S. J., Leahy, R., Lieberman, J. R., Belldegrun, A. S., Sawyers, C. L., Gambhir, S. S. 2002; 29 (9): 1225-1236

  Abstract

  The aims were to optimize reproducibility and establish [(18)F]fluoride ion bone scanning in mice, using a dedicated small animal positron emission tomography (PET) scanner (microPET) and to correlate functional findings with anatomical imaging using computed tomography (microCAT). Optimal tracer uptake time for [(18)F]fluoride ion was determined by performing dynamic microPET scans. Quantitative reproducibility was measured using region of interest (ROI)-based counts normalized to (a) the injected dose, (b) integral of the heart time-activity curve, or (c) ROI over the whole skeleton. Bone lesions were repetitively imaged. Functional images were correlated with X-ray and microCAT. The plateau of [(18)F]fluoride uptake occurs 60 min after injection. The highest reproducibility was achieved by normalizing to an ROI over the whole skeleton, with a mean percent coefficient of variation [(SD/mean) x 100] of <15%-20%. Benign and malignant bone lesions were successfully repetitively imaged. Preliminary correlation of microPET with microCAT demonstrated the high sensitivity of microPET and the ability of microCAT to detect small osteolytic lesions. Whole-body [(18)F]fluoride ion bone imaging using microPET is reproducible and can be used to serially monitor normal and pathological changes to the mouse skeleton. Morphological imaging with microCAT is useful to display correlative changes in anatomy. Detailed in vivo studies of the murine skeleton in various small animal models of bone diseases should now be possible.

  View details for DOI 10.1007/s00259-002-0850-1

  View details for Web of Science ID 000178150400020

  View details for PubMedID 12418463

• Molecular imaging of cancer with positron emission tomography NATURE REVIEWS CANCER Gambhir, S. S. 2002; 2 (9): 683-693

  Abstract

  The imaging of specific molecular targets that are associated with cancer should allow earlier diagnosis and better management of oncology patients. Positron emission tomography (PET) is a highly sensitive non-invasive technology that is ideally suited for pre-clinical and clinical imaging of cancer biology, in contrast to anatomical approaches. By using radiolabelled tracers, which are injected in non-pharmacological doses, three-dimensional images can be reconstructed by a computer to show the concentration and location(s) of the tracer of interest. PET should become increasingly important in cancer imaging in the next decade.

  View details for DOI 10.1038/nrc882

  View details for Web of Science ID 000180447400014

  View details for PubMedID 12209157

• CL1-SR39: A noninvasive molecular imaging model of prostate cancer suicide gene therapy using positron emission tomography JOURNAL OF UROLOGY Pantuck, A. J., Berger, F., Zisman, A., Nguyen, D., Tso, C. L., Matherly, J., Gambhir, S. S., Belldegrun, A. S. 2002; 168 (3): 1193-1198

  Abstract

  We developed a prostate cancer tumor model capable of being noninvasively imaged using positron emission tomography (PET) based on expression of the herpes simplex virus thymidine kinase (HSV1-tk) reporter gene.The androgen independent, metastatic prostate cancer cell lines CL1 and CL1-GFP were stably transfected with the mutant HSV1-tk gene pcDNA3.1/pCMV-sr39tk, which has increased ability to phosphorylate penciclovir. The presence of the sr39tk gene product was analyzed by Western blot analysis and relative thymidine kinase enzyme activity was assessed by a functional thymidine kinase enzyme activity assay. Subcutaneous and orthotopic CL1 and CL1-SR39 tumor xenografts were established in SCID mice. The ability to image CL1-SR39 was assessed using fluorodeoxyglucose and F-penciclovir ( F-FHBG) micro-PET (a rodent PET scanner). To investigate the systemic distribution of intratumoral sr39tk injections established CL1 tumors were transiently injected with first generation adenoviral vectors carrying the sr39tk gene under control of the strong cytomegalovirus promoter Ad-CMV-HSV1-sr39tk and imaged using micro-PET.Transfection of sr39tk into CL1 cells was successful. CL1-SR39 thymidine kinase enzyme activity was greater than twice the activity of the glioma cell line C6-SR39 control and above the threshold necessary for micro-PET detection. Fluorodeoxyglucose micro-PET in SCID mice was positive for CL1 and CL1-SR39 tumors. Selective micro-PET of subcutaneous CL1-SR39 tumors was done using F-FHBG. Micro-PET imaging after systemic and intratumoral injection of Ad-CMV-HSV1-sr39tk revealed significant systemic transgene leakage with significant hepatic expression of sr39TK protein.Molecular based imaging of sr39tk transfected prostate cancer tumors and adenoviral delivered HSV1-tk suicide gene therapy based on the selective conversion and intracellular trapping of F-FHBG by sr39tk is feasible. Potential applications include noninvasive monitoring of the location, duration and intensity of gene constructs, which may contribute to the safety of clinical gene therapy protocols, and noninvasive imaging of the prostate cancer xenograft response to experimental therapy.

  View details for DOI 10.1079/01.ju.0000026576.46595.61

  View details for Web of Science ID 000177539600086

  View details for PubMedID 12187266

• Visualization of advanced human prostate cancer lesions in living mice by a targeted gene transfer vector and optical imaging NATURE MEDICINE Adams, J. Y., Johnson, M., Sato, M., Berger, F., Gambhir, S. S., Carey, M., Iruela-Arispe, M. L., Wu, L. 2002; 8 (8): 891-896

  Abstract

  Non-invasive imaging and transcriptional targeting can improve the safety of therapeutic approaches in cancer. Here we demonstrate the ability to identify metastases in a human-prostate cancer model, employing a prostate-specific adenovirus vector (AdPSE-BC-luc) and a charge-coupled device-imaging system. AdPSE-BC-luc, which expresses firefly luciferase from an enhanced prostate-specific antigen promoter, restricted expression in the liver but produced robust signals in prostate tumors. In fact, expression was higher in advanced, androgen-independent tumors than in androgen-dependent lesions. Repetitive imaging over a three-week period after AdPSE-BC-luc injection into tumor-bearing mice revealed that the virus could locate and illuminate metastases in the lung and spine. Systemic injection of low doses of AdPSE-BC-luc illuminated lung metastasis. These results demonstrate the potential use of a non-invasive imaging modality in therapeutic and diagnostic strategies to manage prostate cancer.

  View details for DOI 10.1038/nm743

  View details for Web of Science ID 000177200900037

  View details for PubMedID 12134144

• Positron emission tomography imaging of cardiac reporter gene expression in living rats CIRCULATION Wu, J. C., Inubushi, M., Sundaresan, G., Schelbert, H. R., Gambhir, S. S. 2002; 106 (2): 180-183

  Abstract

  Imaging reporter gene expression is useful for noninvasive monitoring of gene therapy. In this study, we imaged cardiac reporter gene expression in living rats using micro positron emission tomography (microPET).Rats (n=10) underwent intramyocardial injection with 1x10(9) pfu of adenovirus carrying cytomegalovirus promoter-driving herpes simplex virus type 1 mutant thymidine kinase (Ad-CMV-HSV1-sr39tk) as PET reporter gene. Control rats (n=4) received 1x10(9) pfu of adenovirus carrying cytomegalovirus promoter-driving firefly luciferase (Ad-CMV-Fluc). On days 2 to 4, microPET images were obtained after a tail vein injection of nitrogen-13 ammonia ([13N]-NH3) as myocardial perfusion tracer, followed by 9-(4-[18F]-fluoro-3 hydroxymethylbutyl) guanine ([18F]-FHBG) to assess HSV1-sr39tk expression. After imaging, hearts were removed for ex vivo [18F] gamma counting and thymidine kinase enzyme assay. Results show homogenous myocardial distribution of [13N]-NH3 on all microPET images. Rats injected with Ad-CMV-HSV1-sr39tk have significant [18F]-FHBG uptake in the anterolateral wall compared with background signal in controls. Gamma counting shows 20.0+/-4.4-fold increase of radioactivity, whereas enzyme assay shows 22.1+/-6.1-fold increase of thymidine kinase activity in Ad-CMV-HSV1-sr39tk injected rats (P<0.05).Successful imaging of cardiac HSV1-sr39tk expression was performed in living rats with microPET. The presence of [18F]-FHBG uptake is confirmed by gamma counting and the presence of HSV1-sr39TK protein by thymidine kinase enzyme assay. Cardiac reporter gene imaging by PET may eventually be applied toward human gene therapy studies.

  View details for DOI 10.1161/01.CIR.0000023620.59633.53

  View details for Web of Science ID 000176820500016

  View details for PubMedID 12105155

• Noninvasive, repetitive, quantitative measurement of gene expression from a bicistronic message by positron emission tomography, following gene transfer with adenovirus MOLECULAR THERAPY Liang, Q. W., Gotts, J., Satyamurthy, N., Barrio, J., Phelps, M. E., Gambhir, S. S., Herschman, H. R. 2002; 6 (1): 73-82

  Abstract

  Gene therapy protocols are hampered by the inability to monitor the location, magnitude, and duration of ectopic gene expression following DNA delivery. Consequently, it is difficult to establish quantitative correlations and/or causal relationships between therapeutic gene expression and phenotypic responses in treated individuals. One approach to monitor "therapeutic gene" expression indirectly is to incorporate reporter genes that can be imaged in vivo into bicistronic transcription units, along with the therapeutic genes. Expression of the dopamine D2 receptor (D2R) and herpes simplex virus thymidine kinase (HSV1-TK) can both be monitored, in vivo, by positron-emission tomography (PET). We created ad.DTm, an adenovirus containing a cytomegalovirus (CMV) early promoter-driven transcription unit, in which the D2R gene is placed proximal to an encephalomyocarditis virus internal ribosomal entry site (IRES) and a modified HSV1-tk gene is placed distal to the IRES. Following intravenous ad.DTm injection into mice, correlated hepatic D2R and HSV1-sr39tk PET reporter gene expression was demonstrated. Repeated microPET scanning quantitated both D2R-dependent sequestration of a positron-emitting ligand and HSV1-TK-dependent sequestration of a positron-emitting product. It is possible, in living mice, to investigate noninvasively and to measure quantitatively and repeatedly correlated expression of two coding regions from a bicistronic transcription unit over a 3-month period following adenovirus delivery.

  View details for DOI 10.1006/mthe.2002.0626

  View details for Web of Science ID 000176578500012

  View details for PubMedID 12095306

• Towards in vivo nuclear microscopy: iodine-125 imaging in mice using micro-pinholes EUROPEAN JOURNAL OF NUCLEAR MEDICINE AND MOLECULAR IMAGING Beekman, F. J., McElroy, D. P., Berger, F., Gambhir, S. S., Hoffman, E. J., Cherry, S. R. 2002; 29 (7): 933-938

  Abstract

  Position-sensitive gamma-radiation detectors equipped with collimators have been used for in vivo imaging of the distribution of radiolabelled molecules in laboratory animals and humans for several decades. To date, the best image resolution achieved in a rodent is on the order of 1 mm. Here we demonstrate how a basic and compact gamma camera can be constructed for in vivo radionuclide imaging in small animals, at much higher spatial resolution. Resolution improvements were obtained by combining dense, shaped, micro-pinhole apertures with iodine-125, an isotope with low energy emissions, ease of incorporation into a wide range of molecules, and straightforward translation into the clinic via other isotopes of iodine that are suitable for nuclear medicine imaging. (125)I images of test distributions and a mouse thyroid have been obtained at a resolution of as high as 200 microm using this simple bench-top camera. Possible future applications and extension to ultra-high-resolution emission tomography are discussed.

  View details for DOI 10.1007/s00259-002-0805-6

  View details for Web of Science ID 000177001600014

  View details for PubMedID 12111135

• Added clinical benefit of incorporating 2-deoxy-2-[18F]fluoro-D-glucose with positron emission tomography into the clinical evaluation of patients with cognitive impairment. Molecular imaging and biology Silverman, D. H., Cummings, J. L., Small, G. W., Gambhir, S. S., Chen, W., Czernin, J., Phelps, M. E. 2002; 4 (4): 283-293

  Abstract

  Growing evidence indicates that appropriate incorporation of positron emission tomography (PET) into the evaluation of patients with early symptoms of cognitive decline can improve diagnostic and prognostic accuracy. In the present work, an explicitly defined role for PET and its associated impact on expected clinical outcomes were systematically examined.We compared the relative value of two strategies for assessing whether Alzheimer's disease (AD) was responsible for cognitive decline in geriatric patients, and in subsequently managing those patients according to the recommended standards of the American Academy of Neurology (AAN). The first strategy was based on an approach already endorsed by the AAN, following evidence-based reviews carried out by its quality standards subcommittee. The second approach was based on many of the same AAN recommendations-with respect to initial general medical and neurologic examination, structural imaging and laboratory tests, as well as ultimate management-but additionally incorporated PET in appropriate cases, to determine the presence or absence of a pattern of regional cerebral metabolism characteristic of AD. Clinical outcomes accruing to each strategy were calculated using formalized tools of decision analysis.The strategy making use of PET increased diagnostic accuracy, yielding decreased rates of both false negative (from 8.3 to 3.1%) and false positive (from 23.0 to 11.9%) diagnoses for AD, compared with the conventional strategy. When coupled with AAN treatment recommendations for patients having (or not having) non-severe AD, these differences in diagnostic accuracy corresponded to approximately a 62% decrease in avoidable months of nursing home care, and a 48% decrease in months of unnecessary drug therapy resulting from inaccurate diagnoses. The benefit in clinical outcome of the proposed strategy was maintained over a wide range of values for sensitivity, specificity, and projected impact on need for nursing home care.Use of PET for evaluating early cognitive decline in geriatric patients can add valuable information to the clinical assessment, resulting in a greater number of patients being accurately diagnosed and properly treated. PET can be used to diminish disease-related and treatment-related morbidity of dementia, through earlier institution of appropriate management.

  View details for PubMedID 14537119

• The impact of PET on the management of lung cancer: The referring physician's perspective JOURNAL OF NUCLEAR MEDICINE Seltzer, M. A., Yap, C. S., Silverman, D. H., Meta, J., Schiepers, C., Phelps, M. E., Gambhir, S. S., Rao, J., Valk, P. E., Czernin, J. 2002; 43 (6): 752-756

  Abstract

  (18)F-FDG PET is a molecular whole-body imaging modality that is increasingly being used for diagnosing, staging, and restaging cancer. The objective of this study was to determine referring physicians' perspectives on the impact of (18)F-FDG PET on staging and management of lung cancer.A questionnaire was sent to the 292 referring physicians of 744 consecutive patients with known or suspected lung cancer who were evaluated with PET. Questionnaires on 274 patients were returned (response rate, 37%). Management changes were categorized as intermodality (e.g., surgery to medical, surgery to radiation, and medical to no treatment) or intramodality (e.g., altered medical, surgical, or radiotherapy approach).The primary reasons for PET referral were staging of lung cancer in 61% of patients, diagnosis in 20%, and monitoring of therapy or the course of disease in 6%. Physicians reported that PET caused them to change their decision on clinical stage in 44% of all patients: The disease was upstaged in 29% and downstaged in 15%. PET resulted in intermodality management changes in 39% of patients, whereas 15% had an intramodality change.This survey-based study of referring physicians suggests that PET has a major impact on staging and management of lung cancer.

  View details for Web of Science ID 000176001700011

  View details for PubMedID 12050318

• Positron emission tomography imaging analysis of G2A as a negative modifier of lymphoid leukemogenesis initiated by the BCR-ABL oncogene CANCER CELL Le, L. Q., Kabarowski, J. H., Wong, S., Nguyen, K., Gambhir, S. S., Witte, O. N. 2002; 1 (4): 381-391

  Abstract

  G2A is a lymphocyte-expressed G protein-coupled receptor whose genetic ablation results in the development of autoimmunity. Using HSV-TK reporter gene directed positron emission tomography (PET), we demonstrate that prior to any indication of the onset of illness, mice transplanted with BCR-ABL transduced G2A-deficient bone marrow harbor expanded populations of leukemic cells compared to recipients of wild-type bone marrow. The target cell type and anatomical locations of leukemia development are indistinguishable in animals transplanted with G2A+/+ or G2A-/- cells. Shorter disease latency in the G2A-deficient background is associated with an increased rate of cellular expansion. PET can be successfully applied to the temporal and spatial analysis of Bcr-Abl driven leukemic progression and should have utility for the study of other leukemias and lymphomas.

  View details for Web of Science ID 000178352100012

  View details for PubMedID 12086852

• Optimizing prostate cancer suicide gene therapy using herpes simplex virus thymidine kinase active site variants HUMAN GENE THERAPY Pantuck, A. J., Matherly, J., Zisman, A., Nguyen, D., Berger, F., Gambhir, S. S., Black, M. E., Belldegrun, A., Wu, L. L. 2002; 13 (7): 777-789

  Abstract

  The herpes simplex virus (HSV) thymidine kinase gene (tk) forms the basis of a widely used strategy for suicide gene therapy. A library of HSV thymidine kinase enzyme (TK) active site mutants having different affinities for guanosine analog prodrugs was developed. We sought to determine the optimal combination of tk variant and prodrug specifically for prostate cancer gene therapy, using in vitro and in vivo studies of adenovirally infected CL1, DU-145, and LNCaP tumor lines carrying wild-type tk, tk30, tk75, and sr39tk mutants expressed by a strong, constitutive cytomegalovirus promoter and treated with ganciclovir and acyclovir. In vitro experiments involving prostate cancer (CaP) cell line infection were carried out with a broad range of prodrug concentrations, and cell killing was determined by limiting dilution (colony-forming), MTT, and propidium iodide assays. In vivo studies based on CL1-GFP xenograft experiments were carried out to examine the ability of each TK variant to prevent tumor formation and to inhibit tumor growth and development of metastases in established orthotopic and subcutaneous tumors in SCID mice. Both in vitro and in vivo studies suggest improved killing with the sr39tk variant. Thus, the results suggest that the use of sr39tk in future trials of prostate cancer tk suicide gene therapy may be beneficial.

  View details for Web of Science ID 000175226600001

  View details for PubMedID 11975845

• Optical imaging of cardiac reporter gene expression in living rats CIRCULATION Wu, J. C., Inubushi, M., Sundaresan, G., Schelbert, H. R., Gambhir, S. S. 2002; 105 (14): 1631-1634

  Abstract

  Studies of cardiac gene transfer rely on postmortem analysis using histologic staining or enzyme assays. Noninvasive imaging of the temporal and spatial characteristics of cardiac gene expression in the same subject offers significant advantages.Rats underwent direct myocardial injection via left thoracotomy with adenovirus-expressing firefly luciferase (Ad-CMV-Fluc; n=30). The reporter substrate D-luciferin was injected intraperitoneally. Serial images were acquired by use of a cooled charged couple detector (CCD) camera. Results are expressed as relative light unit per minute (RLU/min). Rats transduced with 1x10(9) plaque-forming units show decremental cardiac luciferase activity over time: 152 070+/-21 170 (day 2), 195 806+/-62 630 (day 5), 7250+/-2941 (day 8), and 2040+/-971 RLU/min (day 14). To assess the detection sensitivity, serially diluted titers of Ad-CMV-Fluc were injected: 1x10(9) (195 393+/-14 896), 1x10(8) (33 777+/-18 179), 1x10(7) (417+/-91), 1x10(6) (185+/-64), 1x10(5) (53+/-1), and control (54+/-1) (P<0.05 for 1x10(9), 1x10(8), and 1x10(7) plaque-forming units versus control adenovirus-expressing mutant thymidine kinase [Ad-CMV-HSV1-sr39tk]; n=3). Finally, rats were euthanized, and in vitro luciferase activity correlated with in vivo CCD signals (r2=0.92).This study demonstrates for the first time the feasibility of imaging the location, magnitude, and time course of cardiac reporter gene expression in living rats. Cardiac gene therapy studies could be aided with wider application of this approach.

  View details for DOI 10.1161/01.CIR.0000014984.95520.AD

  View details for Web of Science ID 000174987300185

  View details for PubMedID 11940538

• Optical Imaging of adenoviral mediated cardiac gene expression in living rats Wu, J. C., Inubushi, M., Schelbert, H., Gambhir, S. S. ELSEVIER SCIENCE INC. 2002: 232A

  View details for Web of Science ID 000174106701037

• Ex vivo cell labeling with Cu-64-pyruvaldehyde-bis(N-4-methylthiosemicarbazone) for imaging cell trafficking in mice with positron-emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Adonai, N., Nguyen, K. N., Walsh, J., Iyer, M., Toyokuni, T., Phelps, M. E., McCarthy, T., MCCARTHY, D. W., Gambhir, S. S. 2002; 99 (5): 3030-3035

  Abstract

  We have used copper-64-pyruvaldehyde-bis(N4-methylthiosemicarbazone) (64Cu-PTSM) to radiolabel cells ex vivo for in vivo positron-emission tomography (PET) imaging studies of cell trafficking in mice and for eventual application in patients. 2-[18F]-Fluoro-2-deoxy-d-glucose (FDG) cell labeling also was evaluated for comparison. 64Cu-PTSM uptake by C6 rat glioma (C6) cells increased for 180 min and then stabilized. The labeling efficiency was directly proportional to 64Cu-PTSM concentration and influenced negatively by serum. Label uptake per cell was greater with 64Cu-PTSM than with FDG. However, both 64Cu-PTSM- and FDG-labeled cells showed efflux of cell activity into supernatant. The 64Cu-PTSM labeling procedure did not interfere significantly with C6 cell viability and proliferation rate. MicroPET images of living mice indicate that tail-vein-injected labeled C6 cells traffic to the lungs and liver. In addition, transient splenic accumulation of radioactivity was clearly detectable in a mouse scanned at 3.33 h postinfusion of 64Cu-PTSM-labeled lymphocytes. In contrast, the liver was the principal organ of tracer localization after tail-vein administration of 64Cu-PTSM alone. These results indicate that in vivo imaging of cell trafficking is possible with 64Cu-PTSM-labeled cells. Given the longer t(1/2) of 64Cu (12.7 h) relative to 18F (110 min), longer cell-tracking periods (up to 24-36 h) should be possible now with PET.

  View details for DOI 10.1073/pnas.052709599

  View details for Web of Science ID 000174284600079

  View details for PubMedID 11867752

• Noninvasive quantitative imaging of protein-protein interactions in living subjects PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Ray, P., Pimenta, H., Paulmurugan, R., Berger, F., Phelps, M. E., Iyer, M., Gambhir, S. S. 2002; 99 (5): 3105-3110

  Abstract

  We are developing methods to image molecular and cellular events in living subjects. In this study, we validate imaging of protein-protein interactions in living mice by using bioluminescent optical imaging. We use the well studied yeast two-hybrid system adapted for mammalian cells and modify it to be inducible. We employ the NF-kappaB promoter to drive expression of two fusion proteins (VP16-MyoD and GAL4-ID). We modulate the NF-kappaB promoter through tumor necrosis factor alpha. Firefly luciferase reporter gene expression is driven by the interaction of MyoD and ID through a transcriptional activation strategy. We demonstrate the ability to detect this induced protein-protein interaction in cell culture and image this induced interaction in living mice by using transiently transfected cells. The current approach will be a valuable and potentially generalizable tool to noninvasively and quantitatively image protein-protein interactions in living subjects. The approaches validated should have important implications for the study of protein-protein interactions in cells maintained in their natural in vivo environment as well as for the in vivo evaluation of new pharmaceuticals targeted to modulate protein-protein interactions.

  View details for DOI 10.1073/pnas.052710999

  View details for Web of Science ID 000174284600092

  View details for PubMedID 11854471

  View details for PubMedCentralID PMC122480

• Molecular engineering of a two-step transcription amplification (TSTA) system for transgene delivery in prostate cancer MOLECULAR THERAPY Zhang, L. Q., Adams, J. Y., Billick, E., Ilagan, R., Iyer, M., Le, K., Smallwood, A., Gambhir, S. S., Carey, M., Wu, L. L. 2002; 5 (3): 223-232

  Abstract

  Gene therapy is founded on the concept that tissue-specific promoters can express heterologous genes for molecular imaging or therapeutic applications. The engineering of cell-specific enhancers to improve potency and the development of two-step transcriptional activation (TSTA) approaches have significantly improved the efficacy of transgene expression. Here we combine these technologies to create a robust, titratable, androgen-responsive system targeted to prostate cancer cells. Our "chimeric" TSTA system uses a duplicated variant of the prostate-specific antigen (PSA) gene enhancer to express GAL4 derivatives fused to one, two, or four VP16 activation domains. We targeted the resulting activators to cells with reporter templates bearing one, two, or five GAL4 binding sites upstream of firefly luciferase. We monitored activity via firefly luciferase assays in transfected cell extracts and in live nude mice using a cooled charge-coupled device (CCD) imaging system. In this system, we found that firefly luciferase expression in prostate cancer cells can be varied over an 800-fold range. We also found that a single plasmid bearing the optimized enhancer, GAL4-VP16 derivative, and reporter expressed firefly luciferase at 20-fold higher levels than the cytomegalovirus enhancer. We discuss the implications of this strategy and its application to molecular imaging and therapy.

  View details for DOI 10.1006/mthe.2002.0551

  View details for Web of Science ID 000174192700005

  View details for PubMedID 11863411

• The impact of 2-deoxy-2[18F] fluoro-D-glucose whole body positron emission tomography for managing patients with melanoma: the referring physician's perspective. Molecular imaging and biology Wong, C., Silverman, D. H., Seltzer, M., Schiepers, C., Ariannejad, M., Gambhir, S. S., Phelps, M. E., Rao, J., Valk, P., Czernin, J. 2002; 4 (2): 185-190

  Abstract

  Whole body positron emission tomography (PET) imaging with 2-deoxy-2[18F]fluoro-D-glucose (FDG) has been used successfully to diagnose and stage melanoma. The impact of FDG-PET, however, on patient stage and management from the referring physicians' perspective is unknown.A questionnaire was sent to referring physicians to investigate whether and how PET altered clinical decision in treatment of melanoma patients. Surveys were sent to referring physicians of every melanoma patient who had a PET scan performed at UCLA or the Northern California PET Imaging Center (NCPIC). Data were used to evaluate the impact of FDG-PET on clinical management of melanoma patients based on pre-PET and post-PET staging. Management changes were classified as inter-modality if therapy changed from one modality to another or intra-modality if changes were made within a treatment modality.Fifty-one questionnaires (response rate of 35%) have been received to date. Referring physicians indicated that whole body FDG-PET changed the clinical stage in 15 out of 51 (29%) patients: 10 (20%) were up-staged and five (10%) were down-staged. The PET findings resulted in inter-modality management changes in 15 out of 51 patients (29%). Intra-modality management change occurred in nine patients (18%).From the referring physicians' perspective, FDG-PET has a major impact and results in management changes in 53% of patients with melanoma.

  View details for PubMedID 14537142

• Gene expression tomography PHYSIOLOGICAL GENOMICS Brown, V. M., Ossadtchi, A., Khan, A. H., Gambhir, S. S., Cherry, S. R., Leahy, R. M., Smith, D. J. 2002; 8 (2): 159-167

  Abstract

  Gene expression tomography, or GET, is a new method to increase the speed of three-dimensional (3-D) gene expression analysis in the brain. The name is evocative of the method's dual foundations in high-throughput gene expression analysis and computerized tomographic image reconstruction, familiar from techniques such as positron emission tomography (PET) and X-ray computerized tomography (CT). In GET, brain slices are taken using a cryostat in conjunction with axial rotation about independent axes to create a series of "views" of the brain. Gene expression information obtained from the axially rotated views can then be used to recreate 3-D gene expression patterns. GET was used to successfully reconstruct images of tyrosine hydroxylase gene expression in the mouse brain, using both RNase protection and real-time quantitative reverse transcription PCR (QRT-PCR). A Monte-Carlo analysis confirmed the good quality of the GET image reconstruction. By speeding acquisition of gene expression patterns, GET may help improve our understanding of the genomics of the brain in both health and disease.

  View details for DOI 10.1152/physiolgenomics.00090.2001

  View details for Web of Science ID 000174252200011

  View details for PubMedID 11875194

• Decision analysis for the cost-effective management of recurrent colorectal cancer. Annals of surgery Park, K. C., Schwimmer, J., Gambhir, S. S. 2002; 235 (2): 309-310

  View details for PubMedID 11807375

  View details for PubMedCentralID PMC1422432

• Evaluating early dementia with and without assessment of regional cerebral metabolism by PET: A comparison of predicted costs and benefits JOURNAL OF NUCLEAR MEDICINE Silverman, D. H., Gambhir, S. S., Huang, H. W., Schwimmer, J., Kim, S., Small, G. W., Chodosh, J., Czernin, J., Phelps, M. E. 2002; 43 (2): 253-266

  Abstract

  Evaluating dementia in patients with early symptoms of cognitive decline is clinically challenging. Growing evidence indicates that appropriate incorporation of PET into the clinical work-up can improve diagnostic and prognostic accuracy with respect to Alzheimer's disease (AD), the most common cause of dementia in the geriatric population. The precise diagnostic role of PET and its economic impact in this context, however, have not been systematically examined previously.We compared the relative value of 2 strategies for assessing whether early AD is responsible for cognitive symptoms in geriatric patients: (a) a conventional approach, based largely on establishing clinical criteria for the presence of dementia and excluding non-AD etiologies that could contribute to the patient's symptoms, and (b) a proposed approach using PET to examine regional cerebral metabolism and look for characteristic patterns of abnormal metabolism. The total costs (measured in dollars) and benefits (measured in number of accurate diagnoses) of diagnostic testing and clinical outcomes accruing to each strategy were calculated using formalized tools of decision analysis. The primary outcome measure by which the strategies were compared was the ratio of costs to benefits obtained following each approach.Following the proposed approach led to improved accuracy in identifying early AD, without adding to the overall costs of diagnosis and treatment ($3,433 vs. $3,564 per patient approached by the proposed or conventional algorithm, respectively). The strategy making use of PET was associated with a reduced rate of false-negative and false-positive findings compared with the conventional approach (3.1% vs. 8.2% and 12.0% vs. 23.0%, respectively, at a prevalence of 51.6% in the studied symptomatic population) and a cost savings of $1,138 per correct diagnosis rendered ($4,047 vs. $5,185). The lower cost per unit benefit for the proposed strategy was maintained over a wide range of tested values for variables of sensitivity, specificity, costs of PET and long-term care, and varying approaches to the use of structural neuroimaging.Appropriate use of PET for evaluating early dementia in geriatric patients can add valuable information to the clinical work-up, without adding to the overall costs of evaluation and management, resulting in a greater number of patients being accurately diagnosed for the same level of financial expenditure. Thus, the opportunity exists for diminishing the morbidity of dementia economically, with earlier institution of more appropriate management in evaluated patients.

  View details for Web of Science ID 000173891900032

  View details for PubMedID 11850493

• Optical imaging of Renilla luciferase reporter gene expression in living mice PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Bhaumik, S., Gambhir, S. S. 2002; 99 (1): 377-382

  Abstract

  Imaging reporter gene expression in living subjects is a rapidly evolving area of molecular imaging research. Studies have validated the use of reporter genes with positron emission tomography (PET), single photon emission computed tomography (SPECT), MRI, fluorescence with wild-type and mutants of green fluorescent protein, as well as bioluminescence using Firefly luciferase enzyme/protein (FL). In the current study, we validate for the first time the ability to image bioluminescence from Renilla luciferase enzyme/protein (RL) by injecting the substrate coelenterazine in living mice. A highly sensitive cooled charge-coupled device camera provides images within a few minutes of photon counting. Cells, transiently expressing the Rluc were imaged while located in the peritoneum, s.c. layer, as well as in the liver and lungs of living mice tail-vein injected with coelenterazine. Furthermore, d-luciferin (a substrate for FL) does not serve as a substrate for RL, and coelenterazine does not serve as a substrate for FL either in cell culture or in living mice. We also show that both Rluc and Fluc expression can be imaged in the same living mouse and that the kinetics of light production are distinct. The approaches validated will have direct applications to various studies where two molecular events need to be tracked, including cell trafficking of two cell populations, two gene therapy vectors, and indirect monitoring of two endogenous genes through the use of two reporter genes.

  View details for Web of Science ID 000173233300068

  View details for PubMedID 11752410

• Welcome to the European Journal of Nuclear Medicine and Molecular Imaging. European journal of nuclear medicine and molecular imaging Gambhir, S. S., Ell, P. J. 2002; 29 (1): 1-2

  View details for PubMedID 11807600

• Radionuclide Imaging of Reporter Gene Expression Brain Mapping: The Methods Sundaresan, G., Gambhir, S. S. edited by Toga, A., Mazziotta, J. C. Academic Press: San Diego . 2002: 799–818
• Monitoring Gene Therapy by Positron Emission Tomography Vector Targeting for Therapeutic Gene Delivery Herschman, H. R., Barrio, J., Satyamurthy, N., Liang, Q., MacLaren, D., Yaghoubi, S., Toyokuni, T., Cherry, S., Phelps, M. E., Gambhir, S. S. edited by Curiel, D., Douglas, J. John Wiley and Sons: New York . 2002: 661–685
• Indirect monitoring of endogenous gene expression by positron emission tomography (PET) imaging of reporter gene expression in transgenic mice. Molecular imaging and biology Green, L. A., Yap, C. S., Nguyen, K., Barrio, J. R., Namavari, M., Satyamurthy, N., Phelps, M. E., Sandgren, E. P., Herschman, H. R., Gambhir, S. S. 2002; 4 (1): 71-81

  Abstract

  Repetitive imaging with microPET of endogenous albumin gene expression by using transgenic mice in which the Herpes Simplex Virus Type 1 thymidine kinase (HSV1-tk) reporter gene is driven by the albumin promoter (AL-HSV1-tk).Transgenic mice were imaged repeatedly on a microPET scanner with approximately 200 microCi of 9-[4-[18F]fluoro-3-(hydroxymethyl)butyl]guanine (FHBG) (a substrate for HSV1-TK enzyme). Four transgenic mice were monitored for body weight, serum albumin, and imaged at the end of each of three dietary phases (17%, 0%, and 25% protein diet). Each phase last 14-21 days. The 0% protein diet has been reported previously to reduce albumin gene expression in rats. Twenty non-transgenic mice of the same strain followed a similar feeding schedule and were monitored for serum albumin, body weight, and sacrificed at various time points for determination of their GAPDH normalized albumin mRNA levels.Transgenic mice showed a relatively high FHBG signal from the liver region as expected. Variation of the mean FHBG signal in two mice with a fixed 17% protein diet over a four-month period was <19% s.d. The mean +/- s.e. FHBG liver standardized uptake value (SUV) in four transgenics went from 4.49 +/- 0.32 to 2.17 +/- 0.52 to 6.21 +/- 0.72 as the mice went through the three diets of 17%, 0%, and 25% sequentially. Non-transgenic mice showed GAPDH normalized albumin mRNA that went from 37.68 +/- 6.04 to 26.41 +/- 4.29 to 52.42 +/- 4.09. The FHBG SUV from transgenics was well correlated with GAPDH normalized albumin mRNA from non-transgenics (r(2) = 0.97) supporting that endogenous gene expression of albumin can be indirectly imaged with FHBG.Measuring correlated changes in albumin expression in wild type mice and HSV1-TK expression by microPET in transgenic mice in which the reporter gene is driven by the albumin promoter demonstrates that the HSV1-tk gene can be used to monitor, in living animals, modulated expression of transgenes.

  View details for PubMedID 14538050

• Two-step transcriptional amplification as a method for imaging reporter gene expression using weak promoters PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Iyer, M., Wu, L., Carey, M., Wang, Y., Smallwood, A., Gambhir, S. S. 2001; 98 (25): 14595-14600

  Abstract

  We are developing assays to image tissue-specific reporter gene expression in living mice by using optical methods and positron emission tomography. Approaches for imaging reporter gene expression depend on robust levels of mRNA and reporter protein. Attempts to image reporter gene expression driven by weak promoters are often hampered by the poor transcriptional activity of such promoters. Most tissue-specific promoters are weak relative to stronger but constitutively expressing viral promoters. In this study, we have validated methods to enhance the transcriptional activity of the prostate-specific antigen promoter for imaging by using a two-step transcriptional amplification (TSTA) system. We used the TSTA system to amplify expression of firefly luciferase (fl) and mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) in a prostate cancer cell line (LNCaP). We demonstrate approximately 50-fold (fl) and approximately 12-fold (HSV1-sr39tk) enhancement by using the two-step approach. The TSTA system is observed to retain tissue selectivity. A cooled charge-coupled device optical imaging system was used to visualize the amplified fl expression in living mice implanted with LNCaP cells transfected ex vivo. These imaging experiments reveal a approximately 5-fold gain in imaging signal by using the TSTA system over the one-step system. The TSTA approach will be a valuable and generalizable tool to amplify and noninvasively image reporter gene expression in living animals by using tissue-specific promoters. The approaches validated should have important implications for study of gene therapy vectors, cell trafficking, transgenic models, as well as studying development of eukaryotic organisms.

  View details for Web of Science ID 000172576900074

  View details for PubMedID 11734653

• Positron emission tomography in evaluation of dementia - Regional brain metabolism and long-term outcome JAMA-JOURNAL OF THE AMERICAN MEDICAL ASSOCIATION Silverman, D. H., Small, G. W., Chang, C. Y., Lu, C. S., de Aburto, M. A., Chen, W., Czernin, J., Rapoport, S. I., Pietrini, P., Alexander, G. E., Schapiro, M. B., Jagust, W. J., Hoffman, J. M., Welsh-Bohmer, K. A., Alavi, A., Clark, C. M., Salmon, E., de Leon, M. J., Mielke, R., Cummings, J. L., Kowell, A. P., Gambhir, S. S., Hoh, C. K., Phelps, M. E. 2001; 286 (17): 2120-2127

  Abstract

  Deficits in cerebral glucose utilization have been identified in patients with cognitive dysfunction attributed to various disease processes, but their prognostic and diagnostic value remains to be defined.To assess the sensitivity and specificity with which cerebral metabolic patterns at a single point in time forecast subsequent documentation of progressive dementia.Positron emission tomography (PET) studies of [(18)F]fluorodeoxyglucose in 146 patients undergoing evaluation for dementia with at least 2 years' follow-up for disease progression at the University of California, Los Angeles, from 1991 to 2000, and PET studies in 138 patients undergoing evaluation for dementia at an international consortium of facilities, with histopathological diagnoses an average of 2.9 years later, conducted from 1984 to 2000.Regional distribution of [(18)F]fluorodeoxyglucose in each patient, classified by criteria established a priori as positive or negative for presence of a progressive neurodegenerative disease in general and of Alzheimer disease (AD) specifically, compared with results of longitudinal or neuropathologic analyses.Progressive dementia was detected by PET with a sensitivity of 93% (191/206) and a specificity of 76% (59/78). Among patients with neuropathologically based diagnoses, PET identified patients with AD and patients with any neurodegenerative disease with a sensitivity of 94% and specificities of 73% and 78%, respectively. The negative likelihood ratio of experiencing a progressive vs nonprogressive course over the several years following a single negative brain PET scan was 0.10 (95% confidence interval, 0.06-0.16), and the initial pattern of cerebral metabolism was significantly associated with the subsequent course of progression overall (P<.001).In patients presenting with cognitive symptoms of dementia, regional brain metabolism was a sensitive indicator of AD and of neurodegenerative disease in general. A negative PET scan indicated that pathologic progression of cognitive impairment during the mean 3-year follow-up was unlikely to occur.

  View details for Web of Science ID 000172032100031

  View details for PubMedID 11694153

• Quantitative imaging of gene induction in living animals GENE THERAPY Sun, X., Annala, A. J., Yaghoubi, S. S., Barrio, J. R., Nguyen, K. N., Toyokuni, T., Satyamurthy, N., Namavari, M., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2001; 8 (20): 1572-1579

  Abstract

  Methods to repeatedly, non-invasively, and quantitatively image gene expression in living animals are rapidly emerging and should fundamentally change studies of gene expression in vivo. We previously developed assays utilizing positron emission tomography (PET) to image reporter gene expression. In this paper we: (1) describe a new bi-directional, tetracycline-inducible system that can be used to pharmacologically induce target gene expression and to quantitatively image induced expression by using a PET reporter gene; (2) demonstrate the potential of this system in transient and stable cell transfection assays; and (3) demonstrate the ability to repetitively and quantitatively image tetracycline and tetracycline analog induction of gene expression in living animals. We utilize the dopamine type-2 receptor (D(2)R) and the mutant herpes-simplex virus type 1 thymidine kinase (HSV1-sr39tk) reporter genes to validate this system. We utilize microPET technology to show that quantitative tomographic imaging of gene induction is possible. We find a high correlation (r(2) = 0.98) between 'target' and reporter gene expression. This work establishes a new technique for imaging time-dependent variation of gene expression both from vectors with inducible promoters and in transgenic animals in which pharmacologic induction of gene expression must be monitored. These techniques may be applied both in gene therapy and for the study of gene expression in transgenic animals.

  View details for Web of Science ID 000172014800008

  View details for PubMedID 11704818

• Noninvasive optical imaging of firefly luciferase reporter gene expression in skeletal muscles of living mice MOLECULAR THERAPY Wu, J. C., Sundaresan, G., Iyer, M., Gambhir, S. S. 2001; 4 (4): 297-306

  Abstract

  The ability to monitor reporter gene expression noninvasively offers significant advantages over current techniques such as postmortem tissue staining or enzyme activity assays. Here we demonstrate a novel method of repetitively tracking in vivo gene expression of firefly luciferase (FL) in skeletal muscles of mice using a cooled charged coupled device (CCD) camera. We first show that the cooled CCD camera provides consistent and reproducible results within +/-8% standard deviation from mean values, and a detection sensitivity (range tested: 1 x 10(4) - 1 x 10(9) plaque form-ing units (pfu)) of 1 x 10(6) pfu of E1-deleted adenovirus expressing FL driven by a cytomegalovirus promoter (Ad-CMV-FL). The duration and magnitude of adenoviral mediated (1 x 10(9) pfu) FL gene expression were then followed over time. FL gene expression in immunocompetent Swiss Webster mice peaks within the first 48 hours, falls by 98% after 20 days, and persists for >150 days. In contrast, FL activity in nude mice remains elevated for >110 days. Finally, transduced Swiss Webster and nude mice were sacrificed to show that the in vivo CCD signals correlate well with in vitro luciferase enzyme assays (r(2)=0.91 and 0.96, respectively). Our findings demonstrate the ability of the cooled CCD camera to sensitively and noninvasively track the location, magnitude, and persistence of FL gene expression. Monitoring of gene therapy studies in small animals may be aided considerably with further extensions of this technique.

  View details for Web of Science ID 000171403200006

  View details for PubMedID 11592831

• Monitoring gene therapy with reporter gene imaging SEMINARS IN NUCLEAR MEDICINE Ray, P., Bauer, E., Iyer, M., Barrio, J. R., Satyamurthy, N., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2001; 31 (4): 312-320

  Abstract

  Rapid advances in imaging technologies and gene transfer strategies offer a great opportunity to optimize clinical trials of human gene therapy. Reporter genes are emerging as very powerful tools to monitor the delivery, magnitude, and time variation of therapeutic gene transfer in vivo. Several reporter genes, such as the herpes simplex virus type 1 thymidine kinase, the dopamine type 2 receptor, and the somatostatin receptor type 2, are currently being successfully used with gamma camera, single photon emission computed tomography, and positron emission tomography imaging. These reporter genes can be coupled with a therapeutic gene of interest to indirectly monitor the expression of the therapeutic gene. Finally, applications of the reporter gene technology to other areas, such as cell trafficking studies and transgenic animal models, are now possible.

  View details for DOI 10.1053/snuc.2001.26209

  View details for Web of Science ID 000172025700010

  View details for PubMedID 11710773

• Noninvasive, quantitative imaging in living animals of a mutant dopamine D2 receptor reporter gene in which ligand binding is uncoupled from signal transduction GENE THERAPY Liang, Q., Satyamurthy, N., Barrio, J. R., Toyokuni, T., Phelps, M. P., Gambhir, S. S., Herschman, H. R. 2001; 8 (19): 1490-1498

  Abstract

  The dopamine D2 receptor (D2R) has been used in adenoviral delivery systems and in tumor cell xenografts as an in vivo reporter gene. D2R reporter gene expression has been non-invasively, repetitively and quantitatively imaged by positron emission tomography (PET), following systemic injection of a positron-labeled ligand (3-(2'-[18F]-fluoroethyl)-spiperone; FESP) and subsequent D2R-dependent sequestration. However, dopamine binding to the D2R can modulate cyclic AMP levels. For optimal utilization of D2R as a reporter gene, it is important to uncouple ligand-binding from Gi-protein-mediated inhibition of cAMP production. Mutation of Asp80 or Ser194 produces D2Rs that still bind [3H]spiperone in transfected cells. The D2R80A mutation completely eliminates the ability of the D2R to suppress forskolin-stimulated cAMP accumulation in response to dopamine, in cells transfected with a D2R80A expression plasmid and in cells infected with replication-defective adenovirus expressing D2R80A. The D2R194A mutation substantially reduces, but does not completely eliminate, dopamine modulation of cAMP levels. Cultured cells infected with adenoviruses expressing D2R and D2R80A demonstrated equivalent [3H]spiperone binding activity. Moreover, hepatic FESP sequestration is equivalent, following intravenous injection of adenoviruses expressing D2R and D2R80A. The D2R80A mutant, which can no longer modulate cAMP levels following ligand binding, has full capability as a PET reporter gene.

  View details for Web of Science ID 000171217000007

  View details for PubMedID 11593362

• Impact of whole-body F-18-FDG PET on staging and managing patients with breast cancer: The referring physician's perspective JOURNAL OF NUCLEAR MEDICINE Yap, C. S., Seltzer, M. A., Schiepers, C., Gambhir, S. S., Rao, J., Phelps, M. E., Valk, P. E., Czernin, J. 2001; 42 (9): 1334-1337

  Abstract

  FDG PET has emerged as an important clinical imaging modality for diagnosing and staging cancer. However, the impact of FDG PET on staging and managing patients with breast cancer from the referring physician's point of view is unknown.The referring physicians of 160 breast cancer patients received standardized questionnaires inquiring if and how PET findings altered their patient's stage and their clinical management decisions. Management changes were classified as intermodality if the change was from one modality to another (e.g., medical to surgical, surgical to radiation, medical to no treatment, and vice versa) or as intramodality if the change was within the same modality (e.g., altered medical or radiotherapy approach).Fifty of the 160 surveys were completed (31% response rate). PET changed the clinical stage in 36% of patients (28% upstaged, 8% downstaged) and resulted in intermodality changes in 28% of patients and intramodality changes in 30% of patients.The results of this prospective survey show that FDG PET has a major impact on the management of breast cancer patients, influencing both clinical stage and management in more than 30% of patients.

  View details for Web of Science ID 000170841000018

  View details for PubMedID 11535721

• Human pharmacokinetic and dosimetry studies of [F-18]FHBG: A reporter probe for imaging herpes simplex virus type-1 thymidine kinase reporter gene expression JOURNAL OF NUCLEAR MEDICINE Yaghoubi, S., Barrio, J. R., Dahlbom, M., Iyer, M., Namavari, M., Goldman, R., Herschman, H. R., Phelps, M. E., Gambhir, S. S. 2001; 42 (8): 1225-1234

  Abstract

  9-[4-[(18)F]fluoro-3-(hydroxymethyl)butyl]guanine ([(18)F]FHBG) has been used as a reporter probe to image expression of herpes simplex virus type-1 thymidine kinase (HSV1-tk) reporter gene in living animals. Our aim was to study the kinetics, biodistribution, stability, dosimetry, and safety of [(18)F]FHBG in healthy human volunteers, preparatory to imaging patients undergoing HSV1-tk gene therapy.[(18)F]FHBG was synthesized with a specific activity of 37,000--444,000 GBq/mmol and a radiochemical purity > 99%. Ten healthy volunteers consented to participate in the study. A transmission scan was obtained before bolus injection of 70.3--229.4 MBq [(18)F]FHBG into a hand vein, followed by dynamic PET imaging with 4 consecutive emission scans. Warmed hand-vein blood was withdrawn at various times after injection for blood time--activity measurements. Electrocardiography, blood pressure, and blood and urine pharmacologic parameters were measured before and after injection of the [(18)F]FHBG tracer (n = 5). The stability of [(18)F]FHBG in the urine was analyzed. Attenuation-corrected images were reconstructed using the ordered-subsets expectation maximization algorithm. Image region-of-interest time-activity data were used with the MIRD program to estimate absorbed radiation dosages.[(18)F]FHBG had rapid blood clearance; only 8.42% +/- 4.76% (mean +/- SD) of the peak blood activity remained at approximately 30 min. The average ratio of plasma activity to whole-blood activity during the study was 0.91 +/- 0.04. Penetration of [(18)F]FHBG across the blood-brain barrier was not observed. The primary routes of clearance were renal and hepatobiliary. High activities were observed in the bladder, gut, liver, and kidneys, but <0.0002% of the injected dose per gram was observed in other tissues. In the urine, 83% of activity 180 min after injection was stable [(18)F]FHBG. Blood and urine pharmacologic parameters did not change significantly after injection of the [(18)F]FHBG tracer. The bladder absorbed the highest radiation dose.[(18)F]FHBG has the desirable in vivo characteristics of stability, rapid blood clearance, low background signal, biosafety, and acceptable radiation dosimetry in humans. This study forms the foundation for using [(18)F]FHBG in applications to monitor HSV1-tk reporter gene expression.

  View details for Web of Science ID 000170311300017

  View details for PubMedID 11483684

• Optimization of [18F]Fluoride ion whole body skeletal imaging in mice utilizing microPET and microCAT Berger, F., Loening, A., Chatziioannou, A., Zisman, A., Tso, C., Yoneda, T., Lieberman, J., Sawyers, C., Gambhir, S. SPRINGER-VERLAG. 2001: 1192

  View details for Web of Science ID 000170528300893

• Direct correlation between positron emission tomographic images of two reporter genes delivered by two distinct adenoviral vectors GENE THERAPY Yaghoubi, S. S., Wu, L., Liang, Q., Toyokuni, T., Barrio, J. R., Namavari, M., Satyamurthy, N., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2001; 8 (14): 1072-1080

  Abstract

  Biodistribution, magnitude and duration of a therapeutic transgene's expression may be assessed by linking it to the expression of a positron emission tomography (PET) reporter gene (PRG) and then imaging the PRG's expression by a PET reporter probe (PRP) in living animals. We validate the simple approach of co-administering two distinct but otherwise identical adenoviruses, one expressing a therapeutic transgene and the other expressing the PRG, to track the therapeutic gene's expression. Two PET reporter genes, a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) and dopamine-2 receptor (D(2)R), each regulated by the same cytomegalovirus (CMV) promoter, have been inserted into separate adenoviral vectors (Ad). We demonstrate that cells co-infected with equivalent titers of Ad-CMV-HSV1-sr39tk and Ad-CMV-D(2)R express both reporter genes with good correlation (r(2) = 0.93). Similarly, a high correlation (r(2) = 0.97) was observed between the expression of both PRGs in the livers of mice co-infected via tail-vein injection with equivalent titers of these two adenoviruses. Finally, microPET imaging of HSV1-sr39tk and D(2)R expression with 9-(4-[(18)F]fluoro-3-hydroxymethylbutyl) guanine ([(18)F]FHBG) and 3-(2-[(18)F]fluoroethyl)spiperone ([(18)F]FESP), utilizing several adenovirus-mediated delivery routes, illustrates the feasibility of evaluating relative levels of transgene expression in living animals, using this approach.

  View details for Web of Science ID 000170288800004

  View details for PubMedID 11526454

• A fast nonlinear method for parametric imaging of myocardial perfusion by dynamic N-13-ammonia PET JOURNAL OF NUCLEAR MEDICINE Golish, S. R., Hove, J. D., Schelbert, H. R., Gambhir, S. S. 2001; 42 (6): 924-931

  Abstract

  A parametric image of myocardial perfusion (mL/min/g) is a quantitative image generated by fitting a tracer kinetic model to dynamic (13)N-ammonia PET data on a pixel-by-pixel basis. There are several methods for such parameter estimation problems, including weighted nonlinear regression (WNLR) and a fast linearizing method known as Patlak analysis. Previous work showed that sigmoidal networks can be used for parameter estimation of mono- and biexponential models. The method used in this study is a hybrid of WNLR and sigmoidal networks called nonlinear regression estimation (NRE). The purpose of the study is to compare NRE with WNLR and Patlak analysis for parametric imaging of perfusion in the canine heart by (13)N-ammonia PET.A simulation study measured the statistical performance of NRE, WNLR, and Patlak analysis for a probabilistic model of time-activity curves. Four canine subjects were injected with 740 MBq (13)N-ammonia and scanned dynamically. Images were reconstructed with filtered backprojection and resliced into short-axis cuts. Parametric images of a single midventricular plane per subject were generated by NRE, WNLR, and Patlak analysis. Small regions of interest (ROIs) were drawn on each parametric image (8 ROIs per subject for a total of 32).For the simulation study, the median absolute value of the relative error for a perfusion value of 1.0 mL/min/g was 16.6% for NRE, 17.9% for WNLR, 19.5% for Patlak analysis, and 14.5% for an optimal WNLR method (computable by simulation only). All methods are unbiased conditioned on a wide range of perfusion values. For the canine studies, the least squares line fits comparing NRE (y) and Patlak analysis (z) with WNLR (x) for all 32 ROIs were y = 1.02x - 0.028 and z = 0.90x + 0.019, respectively. Both NRE and Patlak analysis generate 128 x 128 parametric images in seconds.The statistical performance of NRE is competitive with WNLR and superior to Patlak analysis for parametric imaging of myocardial perfusion. NRE is a fast nonlinear alternative to Patlak analysis and other fast linearizing methods for parametric imaging. NRE should be applicable to many other tracers and tracer kinetic models.

  View details for Web of Science ID 000169143000021

  View details for PubMedID 11390558

• A tabulated summary of the FDG PET literature JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Czernin, J., Schwimmer, J., Silverman, D. H., Coleman, R. E., Phelps, M. E. 2001; 42 (5): 1S-93S

  View details for Web of Science ID 000168669400001

  View details for PubMedID 11483694

• Impact of F-18-FDG PET on managing patients with colorectal cancer: The referring physician's perspective JOURNAL OF NUCLEAR MEDICINE Meta, J., Seltzer, M., Schiepers, C., Silverman, D. H., Ariannejad, M., Gambhir, S. S., Phelps, M. E., Valk, P., Czernin, J. 2001; 42 (4): 586-590

  Abstract

  Whole-body PET imaging with 18F-FDG has been used successfully to stage colorectal cancer. However, the impact of FDG PET on patient management from the referring physician's point of view has not been determined.A questionnaire was sent to referring physicians to determine whether and how PET altered the management of colorectal cancer patients. Management changes, when present, were classified as intermodality (e.g., medical to surgical, surgical to radiation, medical to no treatment) or intramodality (e.g., altered medical, surgical, or radiotherapy approach).Of 60 responses from referring physicians, changes in clinical stage were reported for 25 patients (42%). Among these, the disease was upstaged in 20 patients (80%) and downstaged in 5 patients (20%). The PET findings contributed to intermodality management changes in 22 of the 60 patients (37%), intramodality changes in 11 patients (18%), a combination of management changes in 4 patients (7%), and no change in 19 patients (32%). Two of the 60 patients (3%) had other changes, and no response to this question was received for the remaining 2 patients (3%). As a result of PET findings, physicians avoided major surgery in 41% of patients for whom surgery was the intended treatment.This survey-based study of referring physicians shows that FDG PET had a major impact on the management of colorectal cancer patients and contributed to changes in clinical stage and major management decisions in >40% of patients.

  View details for Web of Science ID 000168000500013

  View details for PubMedID 11337546

• Decision Analysis for the Cost-Effective Management of Recurrent Colorectal Cancer. Annals of Surgery Park KC, Schwimmer J, Shepherd JE, Phelps ME, Czernin JR, Schiepers C, Gambhir SS 2001; 233(3): 310-319
• Non-Invasive Imaging of Renilla Luciferase Reporter Gene Expression in Living Mice Academy for Molecular Imaging Bhaumik, S., Gambhir, S. S. 2001
• Comparison of Helical Computerized Tomography, Positron Emission Tomography and Monoclonal Antibody Scans for Evaluation of Lymph Node Metastases in Patients with Prostate Specific Antigen Relapse after Treatment for Localized Prostate Cancer. Journal of Urology Seltzer M, Barbaric Z, Belldegrun A, Naitoh J, Dorey F, Phelps ME, Gambhir SS, Hoh C. 2001; 162(4): 1322-1328
• Bar Harbor Panel on Molecular Imaging. Journal of Nuclear Cardiology Zaret B, Leppo J, Bender J, Carrio I, Echelman W, Gambhir SS, Gree A, Narula J. 2001; 8(2): 256-266
• Use of positron emission tomography in animal research. ILAR journal Cherry, S. R., Gambhir, S. S. 2001; 42 (3): 219-232

  Abstract

  Among the several imaging technologies applied to in vivo studies of research animals, positron emission tomography (PET) is a nuclear imaging technique that permits the spatial and temporal distribution of compounds labeled with a positron-emitting radionuclide to be determined noninvasively. It can be viewed as an in vivo analog of classic autoradiographic methods. Many different positron-labeled compounds have been synthesized as tracers that target a range of specific markers or pathways. These tracers permit the measurement of quantities of biological interest ranging from glucose metabolism to gene expression. PET has been extensively used in imaging studies of larger research animals such as dogs and nonhuman primates. Now, using newly developed high-resolution dedicated animal PET scanners, these types of studies can be performed in small laboratory animals such as mice and rats. The entire whole-body biodistribution kinetics can be determined in a single imaging study in a single animal. This technique should enable statistically significant biodistribution data to be obtained from a handful of animals, compared with the tens or hundreds of animals that might be required for a similar study by autoradiography. PET also enables repeat studies in a single subject, facilitating longitudinal study designs and permitting each animal to serve as its own control in experiments designed to evaluate the effects of a particular interventional strategy. This paper provides a basic overview of the methodology of PET imaging, a discussion of the advantages and drawbacks of PET as a tool in animal research, a description of the latest generation of dedicated animal PET scanners, and a review of a few of the many applications of PET in animal research to date.

  View details for PubMedID 11406721

• Recent advances in imaging endogenous or transferred gene expression utilizing radionuclide technologies in living subjects: applications to breast cancer BREAST CANCER RESEARCH Berger, F., Gambhir, S. S. 2001; 3 (1): 28-U1

  Abstract

  A variety of imaging technologies is being investigated as tools for studying gene expression in living subjects. Two technologies that use radiolabeled isotopes are single photon emission computed tomography (SPECT) and positron emission tomography (PET). A relatively high sensitivity, a full quantitative tomographic capability, and the ability to extend small animal imaging assays directly into human applications characterize radionuclide approaches. Various radiolabeled probes (tracers) can be synthesized to target specific molecules present in breast cancer cells. These include antibodies or ligands to target cell surface receptors, substrates for intracellular enzymes, antisense oligodeoxynucleotide probes for targeting mRNA, probes for targeting intracellular receptors, and probes for genes transferred into the cell. We briefly discuss each of these imaging approaches and focus in detail on imaging reporter genes. In a PET reporter gene system for in vivo reporter gene imaging, the protein products of the reporter genes sequester positron emitting reporter probes. PET subsequently measures the PET reporter gene dependent sequestration of the PET reporter probe in living animals. We describe and review reporter gene approaches using the herpes simplex type 1 virus thymidine kinase and the dopamine type 2 receptor genes. Application of the reporter gene approach to animal models for breast cancer is discussed. Prospects for future applications of the transgene imaging technology in human gene therapy are also discussed. Both SPECT and PET provide unique opportunities to study animal models of breast cancer with direct application to human imaging. Continued development of new technology, probes and assays should help in the better understanding of basic breast cancer biology and in the improved management of breast cancer patients.

  View details for Web of Science ID 000167344300008

  View details for PubMedID 11250742

• 8-[F-18]fluoropenciclovir: An improved reporter probe for imaging HSV1-tk reporter gene expression in vivo using PET JOURNAL OF NUCLEAR MEDICINE Iyer, M., Barrio, J. R., Namavari, M., Bauer, E., Satyamurthy, N., Nguyen, K., Toyokuni, T., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2001; 42 (1): 96-105

  Abstract

  We have synthesized and evaluated 8-[18F]fluoropenciclovir (FPCV) and compared it with 8-[18F]fluoroganciclovir (FGCV) for monitoring the expression of herpes simplex virus type 1 thymidine kinase (HSV1 -tk) reporter gene in cell culture and in vivo.C6 rat glioma cells stably transfected with HSV1-tk (C6-stb-tk+) and control C6 cells were evaluated for their ability to accumulate FGCV versus FPCV. For in vivo studies, 15 mice were injected by tail vein with increasing levels of an adenoviral vector carrying HSV1-tk. Forty-eight hours later the mice were injected with FPCV and killed 3 h later. The percentage injected dose per gram (%ID/g) liver was then determined. Two additional mice were studied by microPET and autoradiography using FPCV to image adenoviral-mediated hepatic HSV1-tk reporter gene expression. A tumor-bearing mouse (C6 control and C6-stb-tk+) was imaged with FDG, FGCV, and FPCV. Two mice carrying tumors expressing two different reporter genes, HSV1-tk and dopamine type 2 receptor (D2R), were also imaged by microPET using FPCV (day 1) and 3-(2'-[18F]fluoroethyl)spiperone (FESP) (day 2).FPCV shows a significantly greater accumulation in C6-stb-tk+ cells than does FGCV (P < 0.05). Over identical ranges of adenoviral administration, mouse liver shows a higher %ID/g liver for FPCV (0%-9%) compared with our previously reported results with FGCV (0%-3%). In C6 control and C6-stb-tk+ tumor-bearing mice, FPCV has a greater accumulation than does FGCV for equal levels of HSV1-tk gene expression. In mice carrying tumors expressing either HSV1-tk or D2R reporter genes, there is a corresponding retention of FPCV and FESP, respectively.These results indicate that FPCV is a better reporter probe than is FGCV for imaging lower levels of HSV1 -tk gene expression in vivo. The results also reveal the ability to monitor the expression of two distinct reporter genes in the same animal using reporter probes specific for each gene.

  View details for Web of Science ID 000166429400038

  View details for PubMedID 11197989

• PET imaging of transgene expression BIOLOGICAL PSYCHIATRY MacLaren, D. C., Toyokuni, T., Cherry, S. R., Barrio, J. R., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2000; 48 (5): 337-348

  Abstract

  A vital step in transgenic animal study and gene therapy is the ability to assay the extent of transgene expression. Unfortunately, classic methods of assaying transgene expression require biopsies or death of the subject. We are developing techniques to noninvasively and repetitively determine the location, duration, and magnitude of transgene expression in living animals. This will allow investigators and clinicians to assay the effectiveness of their particular experimental and therapeutic paradigms. Of radionuclide (single photon emission computed tomography, positron emission tomography [PET]), optical (green fluorescent protein, luciferase), and magnetic (magnetic resonance imaging) approaches, only the radionuclide approach has sufficient sensitivity and quantitation to measure the expression of genes in vivo. We describe the instrumentation involved in high resolution PET scanning. We also describe the principles of PET reporter gene/reporter probe in vivo imaging, the development of two in vivo reporter gene imaging systems, and the validation of our ability to noninvasively, quantitatively, and repetitively image gene expression in murine viral gene transfer and transgenic models. We compare the two reporter gene systems and discuss their utility for the study of transgenic animals and gene therapies. Finally, we mention alternative approaches to image gene expression by using radiolabeled antibody fragments to image specific proteins and radiolabeled oligonucleotides to image RNA messages directly.

  View details for Web of Science ID 000089232700001

  View details for PubMedID 10978717

• Quantification of target gene expression by imaging reporter gene expression in living animals NATURE MEDICINE Yu, Y. J., Annala, A. J., Barrio, J. R., Toyokuni, T., Satyamurthy, N., Namavari, M., Cherry, S. R., Phelps, M. E., Herschman, H. R., Gambhir, S. S. 2000; 6 (8): 933-937

  View details for Web of Science ID 000165473800040

  View details for PubMedID 10932234

• Prevalence of myocardial viability as detected by positron emission tomography in patients with ischemic cardiomyopathy - Response CIRCULATION Czernin, J., Auerbach, M. A., Schoder, H., Hoh, C., Gambhir, S. S., Yaghoubi, S., Sayre, J. W., Silverman, D., Phelps, M. E., Schelbert, H. R. 2000; 102 (4): E31-E31

  View details for Web of Science ID 000088374400006

• High-resolution microPET imaging of carcino-embryonic antigen-positive xenografts by using a copper-64-labeled engineered antibody fragment PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Wu, A. M., Yazaki, P. J., Tsai, S. W., Nguyen, K., Anderson, A. L., MCCARTHY, D. W., Welch, M. J., Shively, J. E., Williams, L. E., Raubitschek, A. A., Wong, J. Y., Toyokuni, T., Phelps, M. E., Gambhir, S. S. 2000; 97 (15): 8495-8500

  Abstract

  Rapid imaging by antitumor antibodies has been limited by the prolonged targeting kinetics and clearance of labeled whole antibodies. Genetically engineered fragments with rapid access and high retention in tumor tissue combined with rapid blood clearance are suitable for labeling with short-lived radionuclides, including positron-emitting isotopes for positron-emission tomography (PET). An engineered fragment was developed from the high-affinity anticarcinoembryonic antigen (CEA) monoclonal antibody T84.66. This single-chain variable fragment (Fv)-C(H)3, or minibody, was produced as a bivalent 80 kDa dimer. The macrocyclic chelating agent 1,4,7, 10-tetraazacyclododecane-N,N',N", N"'-tetraacetic acid (DOTA) was conjugated to the anti-CEA minibody for labeling with copper-64, a positron-emitting radionuclide (t(1/2) = 12.7 h). In vivo distribution was evaluated in athymic mice bearing paired LS174T human colon carcinoma (CEA positive) and C6 rat glioma (CEA negative) xenografts. Five hours after injection with (64)Cu-DOTA-minibody, microPET imaging showed high uptake in CEA-positive tumor (17.9% injected dose per gram +/- 3.79) compared with control tumor (6.0% injected dose per gram +/- 1.0). In addition, significant uptake was seen in liver, with low uptake in other tissues. Average target/background ratios relative to neighboring tissue were 3-4:1. Engineered antibody fragments labeled with positron-emitting isotopes such as copper-64 provide a new class of agents for PET imaging of tumors.

  View details for Web of Science ID 000088273900057

  View details for PubMedID 10880576

• A meta-analysis of the literature for whole-body FDG PET detection of recurrent colorectal cancer JOURNAL OF NUCLEAR MEDICINE Huebner, R. H., Park, K. C., Shepherd, J. E., Schwimmer, J., Czernin, J., Phelps, M. E., Gambhir, S. S. 2000; 41 (7): 1177-1189

  Abstract

  A meta-analysis of the literature for the use of FDG PET in the detection of recurrent colorectal cancer (CRC) was conducted to evaluate the quality of the reported studies. Overall values for the sensitivity and specificity of whole-body FDG PET and an overall FDG PET-directed percentage change in management were also determined through this analysis.Guidelines to evaluate the articles were formulated on the basis of the U.S. medical payer source criteria for assessing studies that report information on usage of new medical technology. A metaanalysis was conducted using methodology described in the peer-reviewed literature.On the basis of the guidelines established for our review, the availability of necessary information for assessing the reliability of the FDG PET data for diagnosing recurrent CRC was less than ideal. Through a meta-analysis of 11 articles, we determined, within a 95% confidence level, an overall sensitivity of 97% (95% confidence level, 95%-99%) and an overall specificity of 76% (95% confidence level, 64%-88%) for FDG PET detecting recurrent CRC throughout the whole body. Furthermore, through pooling of the change-in-management data, an overall FDG PET-directed change in management was calculated to be 29% (95% confidence level, 25%-34%).Our review suggests that improvements can be made to more effectively report the results of these FDG PET studies. The overall values determined through the meta-analysis indicate the potential benefits of using FDG PET as a diagnostic or management tool. Furthermore, these values should prove to be useful to assess the cost-effectiveness of using FDG PET in the management of patients with recurrent CRC.

  View details for Web of Science ID 000089888700021

  View details for PubMedID 10914907

• Decision analysis for the cost effectiveness of Sestamibi Scintimammography in minimizing unnecessary biopsies QUARTERLY JOURNAL OF NUCLEAR MEDICINE Allen, M. W., Hendi, P., Schwimmer, J., Bassett, L., Gambhir, S. S. 2000; 44 (2): 168-185

  Abstract

  The purpose of this study was to assess if breast cancer screening using sestamibi scintimammography (SSMM) in conjunction with mammography (MM) is cost effective in avoiding biopsies in healthy patients.Quantitative decision tree sensitivity analysis was used to compare the conventional MM alone strategy (strategy A) with two decision strategies for screening with SSMM; SSMM after an indeterminate mammogram (strategy B) or SSMM after both a positive and an indeterminate mammogram (strategy C). Cost effectiveness was measured by calculating the expected cost per patient and the average life expectancy per patient for baseline values as well as over a range of values for all of the variables of each strategy.Based on Medicare reimbursement values, strategies B and C showed a cost savings of $9 and $20 per patient respectively as compared to strategy A. This translates into respective savings of $189 and $420 million per year assuming 21 million females undergo screening each year. Strategies B and C did however have a loss of mean life expectancy of 0.000178 and 0.000222 years respectively as compared to strategy A due to interval progression of breast cancer in a small number of women. Strategies B and C significantly lowered the number of biopsies performed on healthy patients in the screening population by 750,063 and 1,557,915 biopsies respectively as compared to strategy A.These results quantitatively verify the potential utility of using SSMM in avoiding unnecessary biopsies.

  View details for Web of Science ID 000088264000007

  View details for PubMedID 10967626

• InternetQuestion and Answer (iQ&A): A Web-based survey technology IEEE TRANSACTIONS ON INFORMATION TECHNOLOGY IN BIOMEDICINE Dennis, R. A., Gambhir, S. S. 2000; 4 (2): 116-125

  Abstract

  This paper presents InternetQuestion and Answer, a Web-based survey development and implementation technology, which has been designed for constructing on-line surveys for educational, medical, or administrative purposes. The system, called iQ&A, is a three-tiered database-backed Web system that has been developed to support a wide range of applications. Surveys are considered as general data collection instruments and include a wide field of application. iQ&A facilitates rapid survey construction and administration which is ideally suited for biomedical research as well as other research and educational activities. Full report management capabilities provide the survey publisher on-line access to current information on survey responses. Current implementations of this technology in the areas of biomedical applications of clinical trials, longitudinal research, and other research-related systems are presented. Further refinement of the current system should lead to a powerful general survey technology for broad-based applications.

  View details for Web of Science ID 000087552400005

  View details for PubMedID 10866410

• A review of the literature for whole-body FDG PET in the management of patients with melanoma QUARTERLY JOURNAL OF NUCLEAR MEDICINE Schwimmer, J., Essner, R., Patel, A., Jahan, S. A., Shepherd, J. E., Park, K., Phelps, M. E., Czernin, J., Gambhir, S. S. 2000; 44 (2): 153-167

  Abstract

  A review and meta-analysis of the literature on the use of 2-[18F]fluoro-2-deoxy-D-glucose (FDG) positron emission tomography (PET) in the detection of recurrent melanoma was conducted. The goals were to evaluate the quality of data reporting and to determine the overall values for the sensitivity and specificity of whole body FDG PET and management changes.Guidelines to evaluate reporting within articles were formulated based on the United States medical payer source criteria for assessing studies reporting information on the utilization of new medical technology. A meta-analysis was conducted using methodology described in the peer reviewed literature.Our MEDLINE PLUS search resulted in a universe of 89 total articles. Within these 89, 19 were categorized in our targeted content area of which 13 were selected for analysis in our targeted subset, with the remaining 70 covering 24 different related content areas. Five of 13 (38%) articles in the target subset reported data which was adequate for incorporation into modeling objectives based on PET sensitivity and specificity values, with 1 of 13 (8%) in the same target subset reporting data adequate for modeling based on change-in-management data. Through a meta-analysis of the 13 target articles we determined, within a 95% confidence level, an overall sensitivity of 92% (95% confidence level 88.41%-95.82%) and an overall specificity of 90% (95% confidence level 83.26%-96.05%) as calculated by number of lesions, for FDG PET detecting recurrent melanoma throughout the whole body. Furthermore, limited data available for change-in-management suggests an overall FDG PET directed change-in-management value of 22%.Our review suggests that improvements can be made to more effectively report the results of these FDG PET studies. The overall values determined through the meta-analysis indicate the potential benefits of using FDG PET as a diagnostic/management tool. Furthermore, these values should prove useful to assessing the cost effectiveness of utilizing FDG PET in the management of recurrent melanoma.

  View details for Web of Science ID 000088264000006

  View details for PubMedID 10967625

• Economic evaluation studies in nuclear medicine: a methodological review of the literature QUARTERLY JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Schwimmer, J. 2000; 44 (2): 121-137

  Abstract

  The growing need for evaluation of the utility of new nuclear medicine technologies has spawned a few economic studies ranging from preliminary indications of cost savings to complete decision analysis models incorporating costs and quality of life. The objective of the current study was to evaluate the methodological quality of economic analyses of nuclear medicine procedures which targeted cost-effectiveness or cost-utility issues published in the medical literature during the years 1985-1999.A computerized literature search was used to identify original investigations from the medical literature which included an economic analysis of a nuclear medicine procedure. Each economic analysis article was evaluated by two independent reviewers for adherence to ten accepted methodological criteria.Of the 29 articles meeting the search criteria, only six (21%) conformed to all ten methodological criteria.Published economic analyses of nuclear medicine procedures usually do not meet accepted methodological standards and could be significantly improved to achieve overall better quality relative to similar analyses in the literature from other medical fields. Continued improvement in the number and quality of economic studies is critically needed for the future competitiveness of nuclear medicine studies.

  View details for Web of Science ID 000088264000004

  View details for PubMedID 10967623

• Economics of nuclear medicine - Introduction QUARTERLY JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 2000; 44 (2): 103-104

  View details for Web of Science ID 000088264000001

  View details for PubMedID 10967620

• Seeing is believing: Non-invasive, quantitative and repetitive imaging of reporter gene expression in living animals, using positron emission tomography JOURNAL OF NEUROSCIENCE RESEARCH Herschman, H. R., MacLaren, D. C., Iyer, M., Namavari, M., Bobinski, K., Green, L. A., Wu, L., Berk, A. J., Toyokuni, T., Barrio, J. R., Cherry, S. R., Phelps, M. E., Sandgren, E. P., Gambhir, S. S. 2000; 59 (6): 699-705

  Abstract

  The ability to monitor reporter gene expression in living animals and in patients will permit longitudinal examinations both of somatically transferred DNA in experimental animals and patients and of transgenic constructs expressed in experimental animals. If investigators can non-invasively monitor the organ and tissue specificity, the magnitude and the duration of gene expression from somatically transferred DNA and from transgenes, conceptually new experimental paradigms will be possible. If clinicians can non-invasively monitor the location, extent and duration of somatically transferred genes, they will be better able to determine the correlations between expression of therapeutic genes and clinical outcomes. We have developed two reporter gene systems for in vivo reporter gene imaging in which the protein products of the reporter genes sequester positron-emitting reporter probes. The "PET reporter gene" dependent sequestration of the "PET reporter probes" is subsequently measured in living animals by Positron Emission Tomography (PET). We describe here the principles of PET reporter gene/PET reporter probe in vivo imaging, the development of two imaging systems, and the validation of their ability to non-invasively, quantitatively and repetitively image reporter gene expression in murine viral gene transfer and transgenic models.

  View details for Web of Science ID 000085698200001

  View details for PubMedID 10700006

• A mutant herpes simplex virus type 1 thymidine kinase reporter gene shows improved sensitivity for imaging reporter gene expression with positron emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Gambhir, S. S., Bauer, E., Black, M. E., Liang, Q. W., Kokoris, M. S., Barrio, J. R., Iyer, M., Namavari, M., Phelps, M. E., Herschman, H. R. 2000; 97 (6): 2785-2790

  Abstract

  We are developing assays for noninvasive, quantitative imaging of reporter genes with positron emission tomography (PET), for application both in animal models and in human gene therapy. We report here a method to improve the detection of lower levels of PET reporter gene expression by utilizing a mutant herpes simplex virus type 1 thymidine kinase (HSV1-sr39tk) as a PET reporter gene. The HSV1-sr39tk mutant was identified from a library of site-directed mutants. Accumulation (net uptake) of the radioactively labeled substrates [8-(3)H]penciclovir ([8-(3)H]PCV), and 8-[(18)F]fluoropenciclovir (FPCV) in C6 rat glioma cells expressing HSV1-sr39tk is increased by a factor of approximately 2.0 when compared with C6 cells expressing wild-type HSV1-tk. The increased imaging sensitivity of HSV1-sr39tk when FPCV is used is also demonstrated in vivo both with tumor cells stably transfected with either HSV1-tk or HSV1-sr39tk, and after hepatic delivery of HSV1-tk or HSV1-sr39tk by using adenoviral vectors. The use of HSV1-sr39tk as a PET reporter gene and FPCV as a PET reporter probe results in significantly enhanced sensitivity for imaging reporter gene expression in vivo.

  View details for Web of Science ID 000085941400069

  View details for PubMedID 10716999

• Synthesis of 8-[F-18]fluoroguanine derivatives: In vivo probes for imaging gene expression with positron emission tomography NUCLEAR MEDICINE AND BIOLOGY Namavari, M., Barrio, J. R., Toyokuni, T., Gambhir, S. S., Cherry, S. R., Herschman, H. R., Phelps, M. E., Satyamurthy, N. 2000; 27 (2): 157-162

  Abstract

  A new method for the preparation of 8-[(18)F]fluoroguanine derivatives based on a direct radiofluorination reaction has been developed. The radiofluorination of ganciclovir (1a) with [(18)F]F(2) was carried out in absolute ethanol in the presence of tetraethylammonium hydroxide at room temperature to give 8-[(18)F]fluoroganciclovir (3a) in an approximately 1% radiochemical yield. Similarly, 8-[(18)F]fluoropenciclovir (3b), 8-[(18)F]fluoroacyclovir (3c), and 8-[(18)F]fluoroguanosine (3d) were synthesized from penciclovir (1b), acyclovir (1c), and guanosine (1d), respectively, using [(18)F]F(2). The structural analyses of the final products (3a, 3b, 3c, and 3d) were carried out after (18)F decay by (1)H, (13)C, and (19)F nuclear magnetic resonance and high resolution mass spectroscopy.

  View details for Web of Science ID 000086680000007

  View details for PubMedID 10773544

• Imaging transgene expression with radionuclide imaging technologies NEOPLASIA Gambhir, S. S., Herschman, H. R., Cherry, S. R., Barrio, J. R., Satyamurthy, N., Toyokuni, T., Phelps, M. E., Larson, S. M., Balatoni, J., Finn, R., Sadelain, M., Tjuvajev, J., Blasberg, R. 2000; 2 (1-2): 118-138

  Abstract

  A variety of imaging technologies are being investigated as tools for studying gene expression in living subjects. Noninvasive, repetitive and quantitative imaging of gene expression will help both to facilitate human gene therapy trials and to allow for the study of animal models of molecular and cellular therapy. Radionuclide approaches using single photon emission computed tomography (SPECT) and positron emission tomography (PET) are the most mature of the current imaging technologies and offer many advantages for imaging gene expression compared to optical and magnetic resonance imaging (MRI)-based approaches. These advantages include relatively high sensitivity, full quantitative capability (for PET), and the ability to extend small animal assays directly into clinical human applications. We describe a PET scanner (microPET) designed specifically for studies of small animals. We review "marker/reporter gene" imaging approaches using the herpes simplex type 1 virus thymidine kinase (HSV1-tk) and the dopamine type 2 receptor (D2R) genes. We describe and contrast several radiolabeled probes that can be used with the HSV1-tk reporter gene both for SPECT and for PET imaging. We also describe the advantages/disadvantages of each of the assays developed and discuss future animal and human applications.

  View details for Web of Science ID 000086519700009

  View details for PubMedID 10933072

• Imaging Gene Expression: Concepts and Future Outlook Diagnostic Nuclear Medicine Gambhir, S. S. edited by Schiepers, C. Springer-Verlag: Berlin. 2000: 253–271
• Progress Toward in Vivo Imaging of Reporter Gene Expression, Using Positron Emission Tomography in Cancer Gene Therapy. ASCO Spring Educational Book (M. Perry, Ed) American Society of Clinical Oncology Herschman HR, Barrio JR, Satyamurthy S, Toyokuni T, Cherry SR, Phelps ME, Gambhir SS. 2000: 169-177
• ROC and localization ROC analyses of lesion detection in whole-body FDG PET: Effects of acquisition mode, attenuation correction and reconstruction algorithm JOURNAL OF NUCLEAR MEDICINE Farquhar, T. H., Llacer, J., Hoh, C. K., Czernin, J., Gambhir, S. S., Seltzer, M. A., Silverman, D. H., Qi, J. Y., Hsu, C. H., Hoffman, E. J. 1999; 40 (12): 2043-2052

  Abstract

  Receiver operating characteristic (ROC) and localization ROC (LROC) studies were performed to compare lesion detection at the borderline of detectability on images reconstructed with two-dimensional filtered backprojection (FBP) without attenuation correction (a common clinical protocol), three-dimensional FBP without attenuation correction, two-dimensional FBP with segmented attenuation correction and a two-dimensional iterative maximum a posteriori (MAP) algorithm using attenuation correction. Lung cancer was the model for the study because of the prominent role of 18F-fluorodeoxyglucose PET in the staging of lung cancer and the importance of lesion detection for staging.Simulated lung cancer lesions were added to two-dimensional and three-dimensional PET data from healthy volunteers. Data were reconstructed using the four methods. Four nuclear medicine physicians evaluated the images. Detection performance with each method was compared using ROC and LROC analysis. Jackknife analysis provided estimates of statistical significance for differences across all readers for the ROC results.ROC and LROC results indicated statistically significant degradation in detection performance with three-dimensional acquisition (average area under ROC curves [Az] 0.51; average area under LROC curves [A(z,LROC)] 0.13) and segmented attenuation correction (average Az 0.59; average Az,LROC 0.29) compared with two-dimensional FBP without attenuation correction (average Az 0.79; average A(z,LROC) 0.54). ROC and LROC results indicated an improvement in detection performance with iterative MAP reconstruction (average Az 0.83; average A(z,LROC) 0.64) compared with two-dimensional FBP reconstruction; this improvement was not statistically significant.Use of segmented attenuation correction or three-dimensional acquisition with FBP reconstruction is not expected to improve detection of lung lesions on whole-body PET images compared with images with two-dimensional FBP without attenuation correction. The potential improvement in detection obtained with an iterative MAP reconstruction method is small compared with that obtained with two-dimensional FBP without attenuation correction.

  View details for Web of Science ID 000084468600018

  View details for PubMedID 10616885

• Comparison of helical computerized tomography, positron emission tomography and monoclonal antibody scans for evaluation of lymph node metastases in patients with prostate specific antigen relapse after treatment for localized prostate cancer JOURNAL OF UROLOGY Seltzer, M. A., Barbaric, Z., Belldegrun, A., Naitoh, J., Dorey, F., Phelps, M. E., Gambhir, S. S., Hoh, C. K. 1999; 162 (4): 1322-1328

  Abstract

  We compare the detection of metastatic disease by helical computerized tomography (CT), positron emission tomography (PET) with F-18 fluorodeoxyglucose and monoclonal antibody scan with 111indium capromab pendetide in patients with an elevated prostate specific antigen (PSA) after treatment for localized prostate cancer.A total of 45 patients with an elevated PSA (median 3.8 ng./ml.) were studied following definitive local therapy with radical prostatectomy in 33, radiation therapy in 9 and cryosurgery in 3. CT of the abdomen and pelvis, and whole body PET were performed in all patients, of whom 21 also underwent monoclonal antibody scan. Lymph nodes 1 cm. in diameter or greater on CT were considered abnormal and were sampled by fine needle aspiration in 12 patients.PET and CT were positive for distant disease in 50% of 22 patients with PSA greater than 4, and in 4 and 17%, respectively, of 23 with PSA less than 4 ng./ml. The detection rate for metastatic disease was similar for CT and PET, and higher overall than that for monoclonal antibody scan. Monoclonal antibody scan was true positive in only 1 of 6 patients, while PET was true positive in 6 of 9 with CT guided fine needle aspiration proved metastases.CT and PET each detected evidence of metastatic disease in 50% of all patients with a high PSA or PSA velocity (greater than 4 ng./ml. or greater than 0.2 ng./ml. per month, respectively). Both techniques are limited for detecting metastatic disease in patients with a low PSA or PSA velocity. Our data suggest that monoclonal antibody scan has a lower detection rate than CT or PET.

  View details for Web of Science ID 000082510700020

  View details for PubMedID 10492189

• Decision analysis in nuclear medicine JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 1999; 40 (9): 1570-1581

  Abstract

  This review focuses primarily on the methodology involved in properly reviewing the literature for performing a meta-analysis and on methods for performing a formal decision analysis using decision trees. Issues related to performing a detailed metaanalysis with consideration of particular issues, including publication bias, verification bias and patient spectrum, are addressed. The importance of collecting conventional measures of test performance (e.g., sensitivity and specificity) and of changes in patient management to model the cost-effectiveness of a management algorithm is detailed. With greater utilization of the techniques discussed in this review, nuclear medicine researchers should be well prepared to compete for the limited resources available in the current health care environment. Furthermore, nuclear medicine physicians will be better prepared to best serve their patients by using only those studies with a proven role in improving patient management.

  View details for Web of Science ID 000082514900028

  View details for PubMedID 10492381

• Assays for noninvasive imaging of reporter gene expression NUCLEAR MEDICINE AND BIOLOGY Gambhir, S. S., Barrio, J. R., Herschman, H. R., Phelps, M. E. 1999; 26 (5): 481-490

  Abstract

  Repeated, noninvasive imaging of reporter gene expression is emerging as a valuable tool for monitoring the expression of genes in animals and humans. Monitoring of organ/cell transplantation in living animals and humans, and the assessment of environmental, behavioral, and pharmacologic modulation of gene expression in transgenic animals should soon be possible. The earliest clinical application is likely to be monitoring human gene therapy in tumors transduced with the herpes simplex virus type 1 thymidine kinase (HSV1-tk) suicide gene. Several candidate assays for imaging reporter gene expression have been studied, utilizing cytosine deaminase (CD), HSV1-tk, and dopamine 2 receptor (D2R) as reporter genes. For the HSV1-tk reporter gene, both uracil nucleoside derivatives (e.g., 5-iodo-2'-fluoro-2'-deoxy-1-beta-D-arabinofuranosyl-5-iodouracil [FIAU] labeled with 124I, 131I) and acycloguanosine derivatives [e.g., 8-[18F]fluoro-9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]guanine (8-[18F]-fluoroganciclovir) ([18F]FGCV), 9-[(3-[18F]fluoro-1-hydroxy-2-propoxy)methyl]guanine ([18F]FHPG)] have been investigated as reporter probes. For the D2R reporter gene, a derivative of spiperone [3-(2'-[18F]-Fluoroethyl)spiperone ([18F]FESP)] has been used with positron emission tomography (PET) imaging. In this review, the principles and specific assays for imaging reporter gene expression are presented and discussed. Specific examples utilizing adenoviral-mediated delivery of a reporter gene as well as tumors expressing reporter genes are discussed.

  View details for Web of Science ID 000081586800001

  View details for PubMedID 10473186

• A study on the cost effectiveness of sestamibi scintimammography for screening women with dense breasts for breast cancer BREAST CANCER RESEARCH AND TREATMENT Allen, M. W., Hendi, P., Bassett, L., Phelps, M. E., Gambhir, S. S. 1999; 55 (3): 243-258

  Abstract

  The potential impact of Sestamibi scintimammography (SSMM) on the cost effective management of women with dense breasts is not known. This study addresses this issue quantitatively by examining the impact of SSMM based screening strategies on the approximately 3,000,000 women over 40 with very dense breasts (DY patterns) without palpable masses and who have had one or more prior mammograms, who undergo routine screening each year. Quantitative decision tree sensitivity analysis was used to compare the conventional mammography (MM) strategy (strategy A), which does not subject patients with negative mammograms to any further examination until their next screening, with two decision strategies for screening with SSMM; SSMM after a negative mammogram (strategy B) or SSMM as the only screening test for women already identified as having dense breasts by a previous mammogram (strategy C). Cost effectiveness was measured by calculating the incremental cost effectiveness ratio (ICER) of strategies B and C, which is the cost of achieving an additional year of life in the screening population by choosing a SSMM based decision strategy rather than the conventional strategy. Strategies B and C reduced the number of false negative diagnoses by 62% and 8%, respectively. The ICER was $632,000 and $3.18M per life year for strategy B and C, respectively. To be cost effective, the pre-test probability of cancer in the study population must be greater than 3% for strategy B or the cost of SSMM must be less than $50 for strategy C. These results show the ICER of an SSMM based breast cancer screening strategy in the management of patients with dense breasts is not currently within the range (approximately $50,000 per year life saved) of other commonly performed medical interventions that are considered cost effective.

  View details for Web of Science ID 000082271900005

  View details for PubMedID 10517169

• Repetitive, non-invasive imaging of the dopamine D-2 receptor as a reporter gene in living animals GENE THERAPY MacLaren, D. C., Gambhir, S. S., Satyamurthy, N., Barrio, J. R., Sharfstein, S., Toyokuni, T., Wu, L., Berk, A. J., Cherry, S. R., Phelps, M. E., Herschman, H. R. 1999; 6 (5): 785-791

  Abstract

  Reporter genes (e.g. beta-galactosidase, chloramphenicol-acetyltransferase, green fluorescent protein, luciferase) play critical roles in investigating mechanisms of gene expression in transgenic animals and in developing gene delivery systems for gene therapy. However, measuring expression of these reporter genes requires biopsy or death. We now report a procedure to image reporter gene expression repetitively and non-invasively in living animals with positron emission tomography (PET), using the dopamine type 2 receptor (D2R) as a reporter gene and 3-(2'-[18F]fluoroethyl)spiperone (FESP) as a reporter probe. We use a viral delivery system to demonstrate the ability of this PET reporter gene/PET reporter probe system to image reporter gene expression following somatic gene transfer. In mice injected intravenously with replication-deficient adenovirus carrying a D2R reporter gene, PET in vivo measures of hepatic [18F] retention are proportional to in vitro measures of hepatic FESP retention, D2R ligand binding and D2R mRNA. We use tumor-forming cells carrying a stably transfected D2R gene to demonstrate imaging of this PET reporter gene/PET reporter probe system in 'tissues'. Tumors expressing the transfected D2R reporter gene retain substantially more FESP than control tumors. The D2R/FESP reporter gene/reporter probe system should be a valuable technique to monitor, in vivo, expression from both gene therapy vectors and transgenes.

  View details for Web of Science ID 000080186700011

  View details for PubMedID 10505102

• Imaging adenoviral-directed reporter gene expression in living animals with positron emission tomography PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Gambhir, S. S., Barrio, J. R., Phelps, M. E., Iyer, M., Namavari, M., Satyamurthy, N., Wu, L., Green, L. A., Bauer, E., MacLaren, D. C., Nguyen, K., Berk, A. J., Cherry, S. R., Herschman, H. R. 1999; 96 (5): 2333-2338

  Abstract

  We are developing quantitative assays to repeatedly and noninvasively image expression of reporter genes in living animals, using positron emission tomography (PET). We synthesized positron-emitting 8-[18F]fluoroganciclovir (FGCV) and demonstrated that this compound is a substrate for the herpes simplex virus 1 thymidine kinase enzyme (HSV1-TK). Using positron-emitting FGCV as a PET reporter probe, we imaged adenovirus-directed hepatic expression of the HSV1-tk reporter gene in living mice. There is a significant positive correlation between the percent injected dose of FGCV retained per gram of liver and the levels of hepatic HSV1-tk reporter gene expression (r2 > 0.80). Over a similar range of HSV1-tk expression in vivo, the percent injected dose retained per gram of liver was 0-23% for ganciclovir and 0-3% for FGCV. Repeated, noninvasive, and quantitative imaging of PET reporter gene expression should be a valuable tool for studies of human gene therapy, of organ/cell transplantation, and of both environmental and behavioral modulation of gene expression in transgenic mice.

  View details for Web of Science ID 000078956600095

  View details for PubMedID 10051642

• Imaging gene expression: Principles and assays JOURNAL OF NUCLEAR CARDIOLOGY Gambhir, S. S., Barrio, J. R., Herschman, H. R., Phelps, M. E. 1999; 6 (2): 219-233

  View details for Web of Science ID 000079762000009

  View details for PubMedID 10327107

• Nonlinear Function Estimation and Other Numerical Methods for Imaging Myocardial Perfusion by N-13 Ammonia PET. Computers in Cardiology. Golish SR, Hove J, Schelbert HR, Gambhir SS 1999; 26: 651-654
• The lag of cerebral hemodynamics with rapidly alternating periodic stimulation: Modeling for functional MRI MAGNETIC RESONANCE IMAGING Hathout, G. M., Gopi, R. K., Bandettini, P., Gambhir, S. S. 1999; 17 (1): 9-20

  Abstract

  A mathematical model that characterizes the response of venous oxygenation to changes in cerebral blood flow (rCBF) and oxygen consumption has been previously presented. We use this model to examine the dampening phenomenon in functional MRI (fMRI) signals with rapidly alternating periodic stimulation bursts. Using a mass balance approach, the equations for an input-output model are derived and solved using Matlab (the Math Works Inc.). Changes in venous oxygenation are related to the results of fMRI experiments using progressively shorter periods of stimulation. An impulse-response function for the model is derived in an attempt to explore the source of the lag in cerebral hemodynamics. Increasing the frequency of stimulation bursts eventually produces a dampening in the fMRI signal. The dampening phenomenon in fMRI signals occurs with stimulation of high frequency on-off alternation. The dynamics of signal dampening, as well as the impulse-response function of a blood oxygen level-dependent model, lend strong indirect support to the hypothesis that blood oxygen level-dependent contrast at the level of the venous blood pool, rather than R1 inflow effects or changes in oxygenation at the level of the capillary bed, underlies the observed signal changes in fMRI.

  View details for Web of Science ID 000077953600002

  View details for PubMedID 9888394

• Prevalence of Myocardial Viability as Detected by Positron Emission Tomography in Patients with Ischemic Cardiomyopathy Circulation Auerbech MA, Schoder H, Hoh C, Gambhir SS, Yaghoubi S, Sayre JW, Silverman D, Phelps ME, Schelbert HR, Czernin J. 1999; 99: 2921-2926
• Cost-effectiveness of FDG-PET for staging non-small cell lung cancer: A decision analysis 34th Annual Meeting of the Society-of-Thoracic-Surgeons Scott, W. J., Shepherd, J., Gambhir, S. S. ELSEVIER SCIENCE INC. 1998: 1876–83

  Abstract

  Preliminary studies have shown that thoracic positron emission tomography (PET) is more accurate than thoracic computed tomography (CT) for the staging of non-small cell lung carcinoma. In the present study the cost-effectiveness, as measured by national Medicare reimbursed costs, and patient life expectancy are used to compare several thoracic PET-based strategies with a conventional thoracic CT-based strategy for preoperative staging.Five decision strategies for selection of potential surgical candidates were compared; thoracic CT alone or four different strategies that use thoracic CT plus thoracic PET. The various paths of each strategy are dependent on numerous variables that were determined from a review of the medical literature. Life expectancy was calculated using the declining exponential approximation of life expectancy and reduced on the basis of procedural morbidity and mortality. Costs were based on national Medicare reimbursed costs. For all possible outcomes of each strategy, the expected cost and projected life expectancy were determined. The effects of changing one or more variables on the expected cost and life expectancy were studied using sensitivity analysis.A strategy that uses PET only after a negative CT study is shown to be a cost-effective alternative to the CT-alone strategy ($25,286 per life-year saved).These results show through rigorous decision tree analysis the potential cost-effectiveness of using thoracic PET in the management of non-small cell lung carcinoma. Greater use of thoracic PET for nonsmall cell lung carcinoma staging is warranted, and further clinical trials should help to validate the analytic results predicted from this study.

  View details for Web of Science ID 000078294900002

  View details for PubMedID 9930463

• Imaging of adenoviral-directed herpes simplex virus type 1 thymidine kinase reporter gene expression in mice with radiolabeled ganciclovir JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Barrio, J. R., Wu, L., Iyer, M., Namavari, M., Satyamurthy, N., Bauer, E., Parrish, C., MacLaren, D. C., Borghei, A. R., Green, L. A., Sharfstein, S., Berk, A. J., Cherry, S. R., Phelps, M. E., Herschman, H. R. 1998; 39 (11): 2003-2011

  Abstract

  We are developing procedures to repeatedly and noninvasively image the expression of transplanted reporter genes in living animals and in patients, using PET. We have investigated the use of the Herpes Simplex Virus type 1 thymidine kinase gene (HSV1-tk) as a reporter gene and [8-14C]-ganciclovir as a reporter probe. HSV1-tk, when expressed, leads to phosphorylation of [8-14C]-ganciclovir. As a result, specific accumulation of phosphorylated [8-14C]-ganciclovir should occur almost exclusively in tissues expressing the HSV1-tk gene.An adenoviral vector was constructed carrying the HSV1-tk gene along with a control vector. C6 rat glioma cells were infected with either viral vector and uptake of [8-3H]-ganciclovir was determined. In addition, 12 mice were injected with varying levels of either viral vector. Adenovirus administration in mice leads primarily to liver infection. Forty-eight hours later the mice were injected with [8-14C]-ganciclovir, and 1 hr later the mice were sacrificed and biodistribution studies performed. Digital whole-body autoradiography also was performed on separate animals. HSV1-tk expression was assayed, using both normalized HSV1-tk mRNA levels and relative HSV1-TK enzyme levels, in both the cell culture and murine studies.Cell culture, murine tissue biodistribution and murine in vivo digital whole-body autoradiography all demonstrate the feasibility of HSV1-tk as a reporter gene and [8-14C]-ganciclovir as an imaging reporter probe. A good correlation (r2 = 0.86) between the [8-14C]-ganciclovir percent injected dose per gram tissue from HSV1-tk positive tissues and HSV1-TK enzyme levels in vivo was found. An initial study in mice with [8-18F]-fluoroganciclovir and microPET imaging supports further investigation of [8-18F]-fluoroganciclovir as a PET reporter probe for imaging HSV1-tk gene expression.These results demonstrate the feasibility of using [8-14C]-ganciclovir as a reporter probe for the HSV1-tk reporter gene, using an in vivo adenoviral mediated gene delivery system in a murine model. The results form the foundation for further investigation of [8-18F]-fluoroganciclovir for noninvasive and repeated imaging of gene expression with PET.

  View details for Web of Science ID 000076784400039

  View details for PubMedID 9829598

• Evaluating tumor biology and oncological disease with positron-emission tomography SEMINARS IN RADIATION ONCOLOGY Silverman, D. H., Hoh, C. K., Seltzer, M. A., Schiepers, C., Cuan, G. S., Gambhir, S. S., Zheng, L., Czernin, J., Phelps, M. E. 1998; 8 (3): 183-196

  Abstract

  The usefulness of positron-emission tomography (PET) for noninvasive assessment of several biological parameters of neoplastic tissue has been reviewed. Numerous radiotracers have been developed, whose particular distribution in the presence of cancer in vivo serves to distinguish medically relevant properties of the tumor cells with which they associate. That distribution is most accurately determined through use of a PET scanner, to localize and quantify the tracer molecules, in which have been incorporated positron-emitting isotopes. These tracers include hypoxia markers, receptor ligands, substrates for enzymatic modification by the products of expression of specific genes, and precursors of protein anabolism and carbohydrate catabolism. In addition, application of PET to evaluation of patients with some particular cancers has been examined, while placing special emphasis on the level of scientific rigor of the evidence underlying conclusions about appropriate use of PET in oncology.

  View details for Web of Science ID 000074767300006

  View details for PubMedID 9634495

• Rapid synthesis of a 5 '-fluorinated oligodeoxy-nucleotide: A model antisense probe for use in imaging with positron emission tomography (PET) BIOORGANIC & MEDICINAL CHEMISTRY LETTERS Pan, D. F., Gambhir, S. S., Toyokuni, T., Iyer, M. R., Acharya, N., Phelps, M. E., Barrio, J. R. 1998; 8 (11): 1317-1320

  Abstract

  5'-Deoxy-5'-fluoro-O4-methylthymidine was synthesized by the reaction of the corresponding 5'-O-tosylate with KF in the presence of Kryptofix [222] and coupled to a 5'-phosphoramidite-activated CPG-bound oligodeoxynucleotide. The sequence of reactions and purifications were accomplished within 4 h, a necessary condition of the development of radiofluorinated antisense oligodeoxynucleotide probe for use with PET.

  View details for Web of Science ID 000074157600010

  View details for PubMedID 9871758

• A new method to estimate parameters of linear compartmental models using artificial neural networks PHYSICS IN MEDICINE AND BIOLOGY Gambhir, S. S., Keppenne, C. L., Banerjee, P. K., Phelps, M. E. 1998; 43 (6): 1659-1678

  Abstract

  At present, the preferred tool for parameter estimation in compartmental analysis is an iterative procedure; weighted nonlinear regression. For a large number of applications, observed data can be fitted to sums of exponentials whose parameters are directly related to the rate constants/coefficients of the compartmental models. Since weighted nonlinear regression often has to be repeated for many different data sets, the process of fitting data from compartmental systems can be very time consuming. Furthermore the minimization routine often converges to a local (as opposed to global) minimum. In this paper, we examine the possibility of using artificial neural networks instead of weighted nonlinear regression in order to estimate model parameters. We train simple feed-forward neural networks to produce as outputs the parameter values of a given model when kinetic data are fed to the networks' input layer. The artificial neural networks produce unbiased estimates and are orders of magnitude faster than regression algorithms. At noise levels typical of many real applications, the neural networks are found to produce lower variance estimates than weighted nonlinear regression in the estimation of parameters from mono- and biexponential models. These results are primarily due to the inability of weighted nonlinear regression to converge. These results establish that artificial neural networks are powerful tools for estimating parameters for simple compartmental models.

  View details for Web of Science ID 000074257600021

  View details for PubMedID 9651032

• Analytical decision model for the cost-effective management of solitary pulmonary nodules JOURNAL OF CLINICAL ONCOLOGY Gambhir, S. S., Shepherd, J. E., Shah, B. D., Hart, E., Hoh, C. K., Valk, P. E., Emi, T., Phelps, M. E. 1998; 16 (6): 2113-2125

  Abstract

  Multiple strategies are currently being used to manage patients who present with indeterminate solitary pulmonary nodules (SPN). We have used decision-analysis models to assess the cost-effectiveness of various strategies for the diagnosis and management of SPN. Four decision strategies were compared: a wait and watch strategy, a surgery strategy, a computed tomography (CT)-based strategy, and a CT-plus-positron emission tomography (PET) strategy. An incremental cost-effectiveness ratio (ICER) was used to compare all strategies to the wait and watch strategy.A CT-plus-PET strategy was the most cost-effective over a large pretest likelihood (probability of having a malignant nodule), with a range of 0.12 to 0.69. Furthermore, within this likelihood range, the potential cost savings of using the CT-plus-PET strategy over the CT strategy ranged from $91 to $2,200 per patient. This translates to a yearly national savings of approximately $62.7 million.Decision-analysis modeling indicates the potential cost-effectiveness of [18F]2-fluoro-2-deoxy-D-glucose (FDG)-PET in the management of SPN. Furthermore, the decision trees developed can be used to model various features of the management of SPN, including modeling the cost-effectiveness of other newly emerging technologies.

  View details for Web of Science ID 000073919200016

  View details for PubMedID 9626211

• Noninvasive methods for quantitating blood time-activity curves from mouse PET images obtained with fluorine-18-fluorodeoxyglucose 43rd Annual Meeting of The Society-for-Nuclear-Medicine Green, L. A., Gambhir, S. S., Srinivasan, A., Banerjee, P. K., Hoh, C. K., Cherry, S. R., Sharfstein, S., Barrio, J. R., Herschman, H. R., Phelps, M. E. SOC NUCLEAR MEDICINE INC. 1998: 729–34

  Abstract

  The mouse model is currently being explored for various applications with PET imaging. Low resolution of current animal scanners relative to mouse size leads to difficulty in quantitating data from mouse PET images. We have, therefore, investigated methods for determining blood time-activity curves (TACs) from mouse PET studies done with fluorine-18-fluorodeoxyglucose (FDG).Eight mice were fasted, the tail vein was injected with 150-300 microCi of FDG and dynamic images were acquired with a CTI/Siemens (Knoxville, TN) animal tomograph for 64.5 min. Concurrently, 11-14 left ventricle (LV) blood samples were drawn directly from the LV chamber. Organ TACs were obtained by drawing circular regions of interest (ROIs) of various sizes on images of the heart, liver and brain. For each mouse, the FDG model parameter K = (K1 x k3)/(k2 + k3) was estimated by a Patlak algorithm with various estimates of the blood TAC and, as a reference tissue TAC, the brain TAC.Most partial-volume-corrected heart ROI TACs overestimated the LV samples. Blood TACs from heart images produced statistically different estimates of K than did the LV samples. The liver image-derived blood TACs yielded estimates of K that were comparable to those yielded by the LV samples. Estimates of K determined with two directly sampled LV points in conjunction with the liver image-derived TAC were not statistically different from the estimates obtained with the LV samples. The size and location of ROIs on images of the liver minimally affected the TACs.We have shown that it is experimentally possible to obtain a blood TAC from mouse studies by repeatedly sampling from the LV. We have also shown that images of the liver can be used to reliably estimate the blood TAC. Future FDG PET studies with the mouse model will benefit from this demonstrated ability to noninvasively quantitate blood TACs directly from FDG PET images.

  View details for Web of Science ID 000072956600042

  View details for PubMedID 9544690

• Dual spillover problem in the myocardial septum with nitrogen-13-ammonia flow quantitation JOURNAL OF NUCLEAR MEDICINE Hove, J. D., Gambhir, S. S., Kofoed, K. F., Kelbaek, H., Schelbert, H. R., Phelps, M. E. 1998; 39 (4): 591-598

  Abstract

  Conventional cardiac PET modeling techniques for [13N]ammonia flow determination do not fully account for the effects of spillover of activity from the right ventricle (RV) onto the activity in the myocardial septum. The purpose of this study was to investigate and to quantitatively account and correct for this effect.Simulations were performed to determine the error introduced by conventional quantitation using septal time-activity curves, which only account for left ventricle (LV) spillover. Furthermore, we explored two separate methods to account for the dual spillover problem: direct estimation of the RV and LV spillover fractions incorporated into the [13N]ammonia model by using the LV and RV input functions in the fit and estimation of the relative dispersion and time shift between the LV and RV input functions by fitting using only the LV input function. The simulated curves were fitted using a two-compartment [13N]ammonia model. Flow estimates from the conventional model and the models including either of the two correction procedures were compared with canine microsphere data.The influence of RV spillover on flow estimation in the septum is determined by several parameters (e.g., dispersion between the RV and LV input function). Depending on the value of these parameters, the septal flow may be underestimated by 0%-30%. The applied methods for correction of the dual spillover problem were comparable and allow for more accurate quantitation in the septum. The canine microsphere data revealed that flow underestimation in the septum is small but significant.Dual spillover in the myocardial septum can introduce significant errors in the estimation of flow by the conventional [13N]ammonia model fitting method, which does not properly account for the RV spillover. Adjusting for the RV spillover in one of the two proposed methods allows for more accurate quantitation of myocardial septal flow with [13N]ammonia PET data.

  View details for Web of Science ID 000072956600013

  View details for PubMedID 9544662

• Clinical Positron Imaging and the Transition Towards an On-Line Electronic Journal. Clinical Positron Imaging. Gambhir SS 1998; 1(1): 71-79
• Positron Emission Tomography in Gynecologic Malignancies. Journal of Gynecologic Techniques Hoh CK, Seltzer MA, Yong TA, Czernin J, Silverman DHS, Gambhir SS, Phelps ME, Karlan BY. 1998; 4(1): 19-24
• Cost-effectiveness analysis in nuclear medicine JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S. 1998; 39 (1): 17N-18N

  View details for Web of Science ID 000071472400003

  View details for PubMedID 9443723

• MD@: A physician-friendly decision analysis tool M D COMPUTING Gambhir, S. S., Gupta, P., Shepherd, J. E., Allen, M. A., Hoh, C., Maddahi, J., Phelps, M. E. 1998; 15 (1): 40-48

  Abstract

  The cost-effective use of medical resources is increasingly important in justifying strategies for medical diagnosis and management. Although some software is available to help with decision analysis, it can be difficult to use these tools for medical applications. We have developed a prototype package for modeling various medical decision strategies, which can be used with a Macintosh or Windows-based personal computer. The system is graphically based, intuitive, and user-friendly. The user constructs decision trees for comparing alternative strategies for diagnosis and management. Selecting blocks from a library, the user sets mean values for variables such as prevalence, sensitivity, specificity, cost, morbidity, and mortality. The system then generates the probabilities of various pathways, using Bayesian analysis, without requiring the user to enter equations. It displays the best strategy in terms of a particular criterion and, when appropriate, performs sensitivity analysis.

  View details for Web of Science ID 000071517500005

  View details for PubMedID 9458662

• Vitamin C crosses the blood-brain barrier in the oxidized form through the glucose transporters JOURNAL OF CLINICAL INVESTIGATION Agus, D. B., Gambhir, S. S., Pardridge, W. M., Spielholz, C., Baselga, J., Vera, J. C., Golde, D. W. 1997; 100 (11): 2842-2848

  Abstract

  Vitamin C concentrations in the brain exceed those in blood by 10-fold. In both tissues, the vitamin is present primarily in the reduced form, ascorbic acid. We identified the chemical form of vitamin C that readily crosses the blood-brain barrier, and the mechanism of this process. Ascorbic acid was not able to cross the blood-brain barrier in our studies. In contrast, the oxidized form of vitamin C, dehydroascorbic acid (oxidized ascorbic acid), readily entered the brain and was retained in the brain tissue in the form of ascorbic acid. Transport of dehydroascorbic acid into the brain was inhibited by d-glucose, but not by l-glucose. The facilitative glucose transporter, GLUT1, is expressed on endothelial cells at the blood-brain barrier, and is responsible for glucose entry into the brain. This study provides evidence showing that GLUT1 also transports dehydroascorbic acid into the brain. The findings define the transport of dehydroascorbic acid by GLUT1 as a mechanism by which the brain acquires vitamin C, and point to the oxidation of ascorbic acid as a potentially important regulatory step in accumulation of the vitamin by the brain. These results have implications for increasing antioxidant potential in the central nervous system.

  View details for Web of Science ID 000071007300026

  View details for PubMedID 9389750

• Effect of intraluminal thrombus on abdominal aortic aneurysm wall stress JOURNAL OF VASCULAR SURGERY Mower, W. R., Quinones, W. J., Gambhir, S. S. 1997; 26 (4): 602-608

  Abstract

  Abdominal aortic aneurysms (AAAs) rupture when the wall stress exceeds the strength of the vascular tissue. Intraluminal thrombus may absorb tension and reduce AAA wall stress. This study was performed to test the hypothesis that intraluminal thrombus can significantly reduce AAA wall stress.AAA wall stresses were determined by axisymmetric finite element analysis. Model AAAs had external diameters ranging from 2.0 to 4.0 cm. Model parameters included: AAA length, 6 cm; wall thickness, 1.5 mm; Poisson's ratio, 0.49; Young's modulus, 1.0 MPa; and luminal pressure, 1.6 x 10(5) dyne/cm2. Stresses were calculated for each model without thrombus, and then were recalculated with thrombus filling 10% of the AAA cavity. Calculations were repeated as thrombus size was increased in 10% increments and as thrombus elastic modulus increased from 0.01 MPa to 1.0 MPa. Maximum wall stresses were compared between models that had intraluminal thrombus and the unmodified models. Stress reduction greater than 25% was considered significant.The maximum stress reduction of 51% occurred when thrombus with elastic modulus of 1.0 MPa filled the entire AAA cavity. Stresses were reduced by only 25% as modulus decreased to 0.2 MPa. Similarly, decreasing thrombus size by 70% resulted in stress reduction of only 28%. Large AAAs experienced greater stress reduction than small AAAs (48% vs 11%).Intraluminal thrombus can significantly reduce AAA wall stress.

  View details for Web of Science ID A1997YE45200008

  View details for PubMedID 9357460

• PET in oncology: Will it replace the other modalities? SEMINARS IN NUCLEAR MEDICINE Hoh, C. K., Schiepers, C., Seltzer, M. A., Gambhir, S. S., Silverman, D. H., Czernin, J., Maddahi, J., Phelps, M. E. 1997; 27 (2): 94-106

  Abstract

  Medical imaging technology is rapidly expanding and the role of each modality is being redefined constantly. PET has been around since the early sixties and gained clinical acceptance in oncology only after an extreme number of scientific publications. Although PET has the unique ability to image biochemical processes in vivo, this ability is not fully used as a clinical imaging tool. In this overview, the role of PET in relation to other tumor imaging modalities will be discussed and the reported results in the literature will be reviewed. In predicting the future of PET, technical improvements of other imaging modalities need to be dealt with. The fundamental physical principles for image formation with computed tomography (CT), ultrasound (US), magnetic resonance imaging (MRI), photon-emission tomography (PET), and single photon emission CT (SPECT) will not change. The potential variety of radiopharmaceuticals which may be developed is unlimited, however, and this provides nuclear imaging techniques with a significant advantage and adaptive features for future biologic imaging. The current applications of PET in oncology have been in characterizing tumor lesions, differentiating recurrent disease from treatment effects, staging tumors, evaluating the extent of disease, and monitoring therapy. The future developments in medicine may use the unique capabilities of PET not only in diagnostic imaging but also in molecular medicine and genetics. The articles discussed in this review were selected from a literature search covering the last 3 years, and in which comparisons of PET with conventional imaging were addressed specifically. PET studies with the glucose analogue fluorine-18-labeled deoxyglucose (FDG) have shown the ability of detecting tumor foci in a variety of histological neoplasms such as thyroid cancer, breast cancer, lymphoma, lung cancer, head and neck carcinoma, colorectal cancer, ovarian carcinoma, and musculoskeletal tumors. Also, the contribution of the whole body PET (WBPET) imaging technique in diagnosis will be discussed. In the current health care environment, a successful imaging technology must not only change medical management but also demonstrate that those changes improve patient outcome.

  View details for Web of Science ID A1997WX00700003

  View details for PubMedID 9144854

• Cost-Effective Selection of Patients for Coronary Angiography Cost-Effective Selection of Patients for Coronary Angiography Maddahi, J., Gambhir, S. S. 1997; 4 (N2 PT2 SUPPS)
• Decision tree sensitivity analysis for cost-effectiveness of FDG-PET in the staging and management of non-small-cell lung carcinoma JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Hoh, C. K., Phelps, M. E., Madar, I., Maddahi, J. 1996; 37 (9): 1428-1436

  Abstract

  Preliminary studies have shown that PET is more accurate than CT for the staging of non-small-cell lung carcinoma (NSCLC). However, the potential effect of PET on the management of these patients and its cost-effectiveness has not been rigorously studied. Thus, we have used decision tree sensitivity analysis to assess the cost-effectiveness of a PET based strategy for staging of NSCLC.Two decision strategies for selection of potential surgical candidates were compared; thoracic CT alone or thoracic CT and thoracic PET. The first decision tree was conservatively constructed by requiring mediastinoscopy (biopsy) to confirm imaging results so that no patient with surgically curable disease would miss the opportunity for surgery in either strategy. A second less conservative tree in which only nonconcordant results are biopsied was also tested. The various paths of each strategy are dependent on numerous parameters which were determined from a review of the medical literature. Life expectancy was calculated using the declining exponential approximation of life expectancy and reduced based on procedural mortality. Costs were based on mean costs at our institution. For all possible outcomes of each strategy, the expected cost and projected life expectancy were determined. The effect of changing one or more parameters on the expected cost and life expectancy were studied using a sensitivity analysis.The CT + PET strategy in the conservative decision tree showed a saving of $1154 per patient without a loss of life expectancy (increase of 2.96 days) as compared to the alternate strategy of CT alone. Both these effects were the result of improved staging of lung carcinoma prior to the decision for surgery. The CT + PET strategy in the less conservative decision tree showed a savings of $2267 per patient but misses 1.7% of potentially operable patients.These results show through rigorous decision tree analysis, the potential cost-effectiveness of using FDG PET in the management of NSCLC. These results form a basis for detailed study of the results obtained from multicenter trials on the accuracy of PET in NSCLC management. Furthermore, the techniques utilized for decision tree analysis have broad range of applicability to the entire field of nuclear medicine.

  View details for Web of Science ID A1996VF79700010

  View details for PubMedID 8790186

• Symbolic Interactive Modeling Package and Learning Environment (SIMPLE), a New Compartmental Modeling Tool. Proceedings of the Society for Computer Simulation, Gambhir, S., Mahoney DK, Turner MS, Wong ATC, Rosenqvist G, Huang SC, Phelps ME. 1996: 173-180
• A QUANTITATIVE PHYSIOLOGICAL MODEL OF BLOOD OXYGENATION FOR FUNCTIONAL MAGNETIC-RESONANCE-IMAGING INVESTIGATIVE RADIOLOGY Hathout, G. M., Gambhir, S. S., Gopi, R. K., KIRLEW, K. A., Choi, Y., So, G., Gozal, D., Harper, R., Lufkin, R. B., Hawkins, R. 1995; 30 (11): 669-682

  Abstract

  Variations in venous deoxyhemoglobin levels in response to neuronal activation represent a complex interplay between focal changes in cerebral blood flow (CBF), cerebral blood volume (CBV), and regional metabolism. The authors present a mathematic model that characterizes the response of venous oxygenation to changes in these variables.Using a mass balance approach, the equations for a simple input-output model are derived and solved using Matlab. Changes in blood oxygenation are related to available results from functional magnetic resonance imaging experiments.Increases in CBF produce declines in oxygen extraction fraction and venous deoxyhemoglobin according to Fick's law, and are quantitatively in agreement with available magnetic resonance and positron-emission tomography data. A flow-volume envelope defines the changes in CBF relative to CBV.It is possible to obtain a quantitative understanding of changes in blood oxygenation and to relate these changes to the observed dynamics of magnetic resonance signal change in the setting of functional stimulation.

  View details for Web of Science ID A1995TE46900007

  View details for PubMedID 8557508

• PARAMETER-ESTIMATION OF CARDIAC GEOMETRY BY ECG-GATED PET IMAGING - VALIDATION USING MAGNETIC-RESONANCE-IMAGING AND ECHOCARDIOGRAPHY JOURNAL OF NUCLEAR MEDICINE Porenta, G., Kuhle, W., Sinha, S., Krivokapich, J., Czernin, J., Gambhir, S. S., Phelps, M. E., Schelbert, H. R. 1995; 36 (6): 1123-1129

  Abstract

  The purpose of this study was to apply and validate a previously developed model-based image analysis technique which derives estimates of regional myocardial wall thickness and the left ventricular radius directly from gated cardiac PET images.In 11 normal volunteers, gated myocardial 18F-deoxyglucose (FDG) images with 16 equal gates spanning the entire cardiac cycle were acquired for 20 min. To improve count statistics and thus image quality, 3 and 5 of 16 gates were summed to obtain systolic and diastolic images. Based on a five-parameter model, radial profiles from systolic and diastolic PET images were fit by nonlinear regression for myocardial wall thickness, left ventricular radius and tracer activities in the blood pool, the myocardial tissue and the extracardiac background. Echocardiography and gated magnetic resonance imaging (MRI) were performed in 11 and 7 volunteers, respectively.We observed a significant (p < 0.001) correlation between measurements obtained by gated PET imaging and the correlative imaging modalities for myocardial wall thickness and left ventricular radius. While good agreement was observed between measurements of average radial shortening, estimates of average wall thickening differed significantly.This model-based analysis offers accurate estimates of regional recovery coefficients directly from gated cardiac PET images and may also prove useful for the assessment of myocardial contractile function. These recovery coefficients are essential for the correction of partial volume effects when quantitative PET studies are performed.

  View details for Web of Science ID A1995RB81600046

  View details for PubMedID 7769438

• Decision Tree Sensitivity Analysis for Cost Effectiveness of FDG PET in the Staging and Management of Non-Small Cell Lung Carcinoma (NSCLC) Journal of Nuclear Medicine Gambhir, S. S., Hoh, C. K., Madar, I., Phelps, M. E., Maddahi, J. 1995
• REGIONAL MYOCARDIAL BLOOD-FLOW AND GLUCOSE-UTILIZATION IN SYMPTOMATIC PATIENTS WITH HYPERTROPHIC CARDIOMYOPATHY CIRCULATION Nienaber, C. A., Gambhir, S. S., Mody, F. V., Ratib, O., Huang, S. C., Phelps, M. E., Schelbert, H. R. 1993; 87 (5): 1580-1590

  Abstract

  Previous studies suggested the presence of myocardial ischemia in symptomatic patients with hypertrophic cardiomyopathy. Positron emission tomography, a technique that can identify metabolic consequences of ischemia in coronary artery disease, permits the noninvasive measurements of regional myocardial blood flow and glucose metabolism. This new quantitative imaging approach should therefore be suitable for detecting a possible enhancement of glucose utilization in myocardium of patients with hypertrophic cardiomyopathy and thus may help to elucidate the pathomechanism of ischemia in this disease.In 13 symptomatic patients with hypertrophic cardiomyopathy, myocardial blood flow and glucose utilization were measured with intravenous N-13-ammonia and F-18 deoxyglucose at rest and, in four patients, again during supine bicycle exercise. At rest, blood flow was significantly lower in hypertrophied than in normal myocardium (0.78 +/- 0.19 versus 0.99 +/- 0.13 mL.min-1.g-1, p < 0.025), whereas rates of glucose utilization were similar (0.88 +/- 0.31 versus 0.87 +/- 0.35 mumol.min-1.g-1). With exercise, blood flow and glucose utilization failed to increase in hypertrophic and normal segments but became more heterogeneously distributed throughout the left ventricular myocardium. Blood flow-metabolism mismatches indicative of myocardial ischemia were noted in three patients at rest and in three of the four patients during exercise and were due to reduced flow in the presence of maintained glucose uptake. The discordance between flow and glucose metabolism in hypertrophied myocardium was significantly more prominent in younger than in older patients.Normal or even elevated rates of glucose utilization and the presence of diminished blood flow in hypertrophied relative to normal myocardium suggest the presence of myocardial ischemia in symptomatic hypertrophic cardiomyopathy. The age dependence of blood flow metabolism disparity suggests differences in the underlying pathophysiology or severity of disease.

  View details for Web of Science ID A1993LB08400019

  View details for PubMedID 8491014

• QUANTIFICATION OF REGIONAL MYOCARDIAL BLOOD-FLOW USING N-13 AMMONIA AND REORIENTED DYNAMIC POSITRON EMISSION TOMOGRAPHIC IMAGING CIRCULATION Kuhle, W. G., Porenta, G., Huang, S. C., Buxton, D., Gambhir, S. S., Hansen, H., Phelps, M. E., Schelbert, H. R. 1992; 86 (3): 1004-1017

  Abstract

  Regional myocardial blood flow has been quantified using transaxial positron emission tomographic (PET) imaging and tracer kinetic modeling. However, the use of transaxial images limits the accuracy of regional partial volume corrections and the localization of the quantified regional flow values. The purpose of the present study was to overcome both problems by calculating regional flows from reoriented short-axis PET images.Twelve experiments were performed in four dogs. 13N-ammonia was injected intravenously while microspheres were administered into the left atrium during baseline, hyperemic, and low-flow conditions. Serial transaxial frames were acquired with a 15-plane PET scanner and reoriented into short-axis frames. The arterial input function and eight regional myocardial tissue activity curves were derived from the reoriented frames. The arterial input functions were corrected for ammonia metabolites, and the myocardial tissue curves were corrected for spillover of activity, partial volume effects, and heterogeneities in the image's spatial resolution introduced during reorientation. Corrections for regional partial volume were based on estimates of the regional myocardial activity thickness derived from reoriented diastolic images of the heart. The myocardial 13N-ammonia kinetics were described with a two-pool compartmental model. Values of regional myocardial blood flow by PET correlated linearly with those by microspheres (slope, 0.94; y intercept, 0.06 ml/min/g; r = 0.93) over a wide range of flows.Regional myocardial blood flow can be measured accurately and noninvasively from serially acquired and reoriented short-axis 13N-ammonia images, thus overcoming limitations inherent to the use of transaxially acquired images and permitting a more complete evaluation of regional blood flows throughout the left ventricular myocardium.

  View details for Web of Science ID A1992JM27000034

  View details for PubMedID 1516170

• USE OF THE ABDOMINAL-AORTA FOR ARTERIAL INPUT FUNCTION DETERMINATION IN HEPATIC AND RENAL PET STUDIES JOURNAL OF NUCLEAR MEDICINE Germano, G., Chen, B. C., Huang, S. C., Gambhir, S. S., Hoffman, E. J., Phelps, M. E. 1992; 33 (4): 613-620

  Abstract

  A method using the activity in the abdominal aorta of human and animal subjects to noninvasively estimate blood-pool input function in dynamic, abdominal PET scans is proposed and validated in this paper. Partial volume effects due to the aorta's dimensions are corrected by a semi-automated algorithm based on the transaxial resolution in the reconstructed images. The technique was validated by comparing PET measurements of abdominal aortic activity to well counter measurements of arterial blood samples (eight canine renal studies) and to PET measurements of left ventricular cavity activity (eight human hepatic studies). In renal studies, correlation analysis of the areas subtended by the two input functions yielded an essentially unitary slope (1.03 +/- 0.09), with high correlation (R2 greater than 0.95, p less than 0.001). In hepatic studies, similar values (0.99 +/- 0.03 and R2 greater than 0.85, p less than 0.001) were found. Correlation of the blood flow estimates based on the two input functions and a two-compartment model produced slopes of 1.07 +/- 0.16 and 1.03 +/- 0.07, and correlations of (R2 greater than 0.98, p less than 0.001) and (R2 greater than 0.97, p less than 0.001) for the renal and hepatic studies, respectively. We conclude that noninvasive, accurate measurements of the arterial input function by dynamic PET imaging are possible and represent a clinically viable alternative to arterial blood sampling.

  View details for Web of Science ID A1992HM08200032

  View details for PubMedID 1552350

• METABOLIC AND FUNCTIONAL RECOVERY OF ISCHEMIC HUMAN MYOCARDIUM AFTER CORONARY ANGIOPLASTY JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY Nienaber, C. A., Brunken, R. C., Sherman, C. T., Yeatman, L. A., Gambhir, S. S., Krivokapich, J., Demer, L. L., Ratib, O., Child, J. S., Phelps, M. E., Schelbert, H. R. 1991; 18 (4): 966-978

  Abstract

  Although revascularization of hypoperfused but metabolically active human myocardium improves segmental function, the temporal relations among restoration of blood flow, normalization of tissue metabolism and recovery of segmental function have not been determined. To examine the effects of coronary angioplasty on 13 asynergic vascular territories in 12 patients, positron emission tomography and two-dimensional echocardiography were performed before and within 72 h of revascularization. Ten patients underwent late echocardiography (67 +/- 19 days) and eight underwent a late positron emission tomographic study (68 +/- 19 days). The extent and severity of abnormalities of wall motion, perfusion and glucose metabolism were expressed as wall motion scores, perfusion defect scores and perfusion-metabolism mismatch scores. Angioplasty significantly increased mean stenosis cross-sectional area (from 0.95 +/- 0.9 to 2.7 +/- 1.4 mm2) and mean cross-sectional luminal diameter (from 0.9 +/- 0.6 to 1.9 +/- 0.5 mm) (both p less than 0.001). Perfusion defect scores in dependent vascular territories improved early after angioplasty (from 116 +/- 166 to 31 +/- 51, p less than 0.002) with no further improvement on the late follow-up study. The mean perfusion-metabolism mismatch score decreased from 159 +/- 175 to 65 +/- 117 early after angioplasty (p less than 0.01) and to 26 +/- 29 at late follow-up (p less than 0.001 vs. before angioplasty; p = NS vs. early after angioplasty). However, absolute rates of glucose utilization remained elevated early after revascularization, normalizing only at late follow-up.(ABSTRACT TRUNCATED AT 250 WORDS)

  View details for Web of Science ID A1991GH89700012

  View details for PubMedID 1894871

• PARAMETRIC IMAGES OF MYOCARDIAL METABOLIC-RATE OF GLUCOSE GENERATED FROM DYNAMIC CARDIAC PET AND 2-[F-18]FLUORO-2-DEOXY-D-GLUCOSE STUDIES JOURNAL OF NUCLEAR MEDICINE Choi, Y., HAWKINS, R. A., Huang, S. C., Gambhir, S. S., Brunken, R. C., Phelps, M. E., Schelbert, H. R. 1991; 32 (4): 733-738

  Abstract

  We describe a method for generating parametric images of the myocardial metabolic rate of glucose (MMRGlc) with positron emission tomography (PET). The method employs serially acquired images of 2-[18F]fluoro-2-deoxy-D-glucose (FDG) uptake and a Patlak graphical analysis of the image data. The arterial input function is derived from images of the left ventricular blood pool calibrated with 18F-plasma measurements. The approach is computationally fast enough to be used in a clinical environment. The MMRGlc parametric images improve myocardial contrast relative to non-parametric images, especially in studies with poor myocardial uptake of FDG. In addition, MMRGlc parametric images consolidate the large amount of data in a dynamic PET study into a clinically usable image set.

  View details for Web of Science ID A1991FG79700031

  View details for PubMedID 2013815

• A QUANTITATIVE INDEX OF REGIONAL BLOOD-FLOW IN CANINE MYOCARDIUM DERIVED NONINVASIVELY WITH N-13 AMMONIA AND DYNAMIC POSITRON EMISSION TOMOGRAPHY JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY Nienaber, C. A., Ratib, O., Gambhir, S. S., Krivokapich, J., Huang, S. C., Phelps, M. E., Schelbert, H. R. 1991; 17 (1): 260-269

  Abstract

  To derive a quantitative index of regional myocardial blood flow, the arterial input function of the flow tracer N-13 ammonia and the regional myocardial N-13 activity concentrations were noninvasively determined in 29 experiments in eight dogs. N-13 ammonia was administered intravenously and cross-sectional images were acquired dynamically using an ECAT III positron emission tomograph with an effective in-plane resolution of 13.46 mm full-width half-maximum. Time-activity curves were derived from the serial images by assigning regions of interest to the left ventricular myocardium and left ventricular blood pool. Tracer net extractions were estimated from the myocardial time-activity concentrations at various times after tracer injection and the integral of the arterial input function. Myocardial blood flow was altered by intravenous dipyridamole, morphine, propranolol and partial or complete occlusion of the left anterior descending coronary artery, and ranged from 9 to 860 ml/min per 100 g. Estimates of tracer net extractions were most accurate when determined from the myocardial N-13 activity concentrations at 60 s divided by the integral of the arterial input function to that time. These estimates correlated with regional myocardial blood flows determined independently by the microsphere technique by y = x (1 - 0.64(e-114/x); SEE = 22.9; r = 0.94). First pass extraction fractions of N-13 ammonia determined noninvasively with this approach declined with higher flows in a nonlinear fashion and were similar to those determined invasively by direct intracoronary N-13 ammonia injections. The findings indicate that an accurate index of regional myocardial blood flow can be obtained noninvasively by high temporal sampling of arterial and myocardial tracer activity concentrations with positron emission tomography. They also provide a basis for the in vivo application of tracer kinetic principles to derive quantitatively and noninvasively regional rates of functional processes in human myocardium.

  View details for Web of Science ID A1991EU23100040

  View details for PubMedID 1987234

• TRACER KINETIC MODELING APPROACHES FOR THE QUANTIFICATION OF HEPATIC-FUNCTION WITH TECHNETIUM-99M DISIDA AND SCINTIGRAPHY JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., HAWKINS, R. A., Huang, S. C., Hall, T. R., Busuttil, R. W., Phelps, M. E. 1989; 30 (9): 1507-1518

  Abstract

  Serial scintigraphic images following injection of [99mTc]iminodiacetic acid compounds such as [99mTc]diisopropyl-iminodiacetic acid (DISIDA) provide qualitative information about liver function. We have investigated approaches for quantitatively describing liver function in terms of the kinetics of DISIDA extraction and excretion by the liver. Several compartmental model configurations were evaluated. A three-compartment model (blood, hepatic parenchyma, intrahepatic bile) was found to fit the data best and was used in conjunction with dynamic image data to obtain estimates of rate constants for liver extraction and excretion of DISIDA, and mean residence time (MRT) of DISIDA in the liver. A noncompartmental approach based on a parametric deconvolution technique was also used to estimate the noncompartmental mean residence time (MRTnc). To assess limitations of the noncompartmental approach, computer simulations were performed using the three-compartment model to generate time-activity curves followed by analysis of these curves by the noncompartmental method. The effect of plasma total bilirubin level on DISIDA uptake and MRT was also investigated. These techniques are readily adaptable to standard nuclear medicine computing facilities, and could be used in the clinical setting to numerically describe serial DISIDA studies (especially in liver transplant patients) efficiently and noninvasively.

  View details for Web of Science ID A1989AP85000012

  View details for PubMedID 2769404

• SIMPLE NONINVASIVE QUANTIFICATION METHOD FOR MEASURING MYOCARDIAL GLUCOSE-UTILIZATION IN HUMANS EMPLOYING POSITRON EMISSION TOMOGRAPHY AND F-18 DEOXYGLUCOSE JOURNAL OF NUCLEAR MEDICINE Gambhir, S. S., Schwaiger, M., Huang, S. C., Krivokapich, J., Schelbert, H. R., Nienaber, C. A., Phelps, M. E. 1989; 30 (3): 359-366

  Abstract

  To estimate regional myocardial glucose utilization (rMGU) with positron emission tomography (PET) and 2-[18F]fluoro-2-deoxy-D-glucose (FDG) in humans, we studied a method which simplifies the experimental procedure and is computationally efficient. This imaging approach uses a blood time-activity curve derived from a region of interest (ROI) drawn over dynamic PET images of the left ventricle (LV), and a Patlak graphic analysis. The spillover of radioactivity from the cardiac chambers to the myocardium is automatically removed by this analysis. Estimates of rMGU were obtained from FDG PET cardiac studies of six normal human subjects. Results from this study indicate that the FDG time-activity curve obtained from the LV ROI matched well with the arterial plasma curve. The rMGU obtained by Patlak graphic analysis was in good agreement with direct curve fitting results (r = 0.90). The average standard error of the estimate of the Patlak rMGU was low (3%). These results demonstrate the practical usefulness of a simplified method for the estimation of rMGU in humans by PET. This approach is noninvasive, computationally fast, and highly suited for developing parametric images of myocardial glucose utilization rate.

  View details for Web of Science ID A1989T600900015

  View details for PubMedID 2786939

• RADIONUCLIDE EVALUATION OF LIVER-TRANSPLANTS SEMINARS IN NUCLEAR MEDICINE HAWKINS, R. A., Hall, T., Gambhir, S. S., Busuttil, R. W., Huang, S. C., Glickman, S., Marciano, D., Brown, R. K., Phelps, M. E. 1988; 18 (3): 199-212

  Abstract

  Orthotopic liver transplantation is now an established technique for treating patients with various forms of end stage liver disease. The number of centers performing the procedure is increasing and, as the number of transplant recipients in the population increases, many institutions performing nuclear medicine studies will be confronted with requests to evaluate these patients. While a variety of radionuclides are proving useful in this evaluation, the 99mTc iminodiacetic acid (IDA) compounds, particularly 99mTc diisopropyl IDA (DISIDA), will probably account for the majority of radionuclide evaluations of these patients because they are well suited to monitor both structural and functional changes of the graft. The primary application of radionuclide studies is focused in the postoperative period, when problems with the vascular and biliary anastomoses, rejection, infections, and bile leaks all produce alterations in radionuclide hepatobiliary studies. Abnormalities such as rejection and infection produce primarily functional, rather than structural changes and are not easily differentiated based upon the kinetics of 99mTc-DISIDA extraction and excretion by the liver, serial imaging and correlation with clinical data is necessary in such situations. Quantitative analyses of kinetic 99mTc IDA (DISIDA) studies and quantitative approaches with other compounds such as 99mTc galactosyl-neoglycoalbumin (NGA) may permit better assessments of relatively subtle changes in liver function in the posttransplant period.

  View details for Web of Science ID A1988P737200003

  View details for PubMedID 3051393

• A STUDY OF THE SINGLE COMPARTMENT TRACER KINETIC-MODEL FOR THE MEASUREMENT OF LOCAL CEREBRAL BLOOD-FLOW USING O-15-WATER AND POSITRON EMISSION TOMOGRAPHY JOURNAL OF CEREBRAL BLOOD FLOW AND METABOLISM Gambhir, S. S., Huang, S. C., HAWKINS, R. A., Phelps, M. E. 1987; 7 (1): 13-20

  Abstract

  The effects of varying the data collection time on the calculation of cerebral blood flow and distribution volume via the integrated projection technique were studied in four human subjects. The significance of these results in terms of the limitations of the single compartment model for 15O-water was explored using computer simulations. The simulations helped to account for causes for the variations seen in blood flow and distribution volume as a function of the data collection time. Two different compartmental models were explored for better quantitation of blood flow and distribution volume.

  View details for Web of Science ID A1987G010000003

  View details for PubMedID 3492506

• Imaging of VEGF Receptor in a Rat Myocardial Infarction Model using Positron Emission Tomography J Nucl Med Rodriguez-Porcel M, Cai W, Gheysens O, Chen I, Chen K, He L, Willman J, Wu JC, Chen X, Gambhir SS : Accepted